Investigation of Rapid Emergence of Colistin Resistance in a Newborn Infected with KPC2-Producing Hypervirulent Carbapenem-Resistant Klebsiella pneumoniae.

OBJECTIVES: Hypervirulent carbapenem-resistant Klebsiella pneumoniae (hv-CRKp) poses a significant threat to public health. This study reports an infection related to hv-CRKp in a premature infant and reveals its colistin resistance and evolutionary mechanisms within the host. METHODS: Three KPC-producing CRKp strains were isolated from a patient with sepsis and CRKp osteoarthritis who had been receiving colistin antimicrobial therapy. The minimum inhibitory concentrations (MICs) of Ceftazidime,Ceftazidime-Avibactam(CAZ-AVI),Meropenem,Imipenem,Tigecycline,Amikacin,Minocycline,Sulfamethoxazole/Trimethoprim,Ciprofloxacin,Levofloxacin,Aztreonam,Cefepime,Cefoperazone/Sulbactam,Piperacillin/Tazobactam and colistin were determined using the microbroth dilution method.The whole-genome sequencing analysis was conducted to determine the STs, virulence genes, and antibiotic resistance genes of three CRKp strains. RESULTS: Whole-genome sequencing revealed that all three CRKp strains belonged to the sequence type (ST) 11 clone and carried a plasmid encoding blaKPC-2. The three strains all possessed the iucABCDiutA virulence cluster, peg-344 gene, and rmpA/rmpA2 genes, defining them as hv-CRKp. Further experiments and whole-genome analysis revealed that a strain of Kp has developed resistance to colistin. The mechanism found to be responsible for the colistin resistance was a deletion mutation of approximately 9000 bp including mgrB gene. CONCLUSION: This study characterizes the colistin resistance of ST11 clone hv-CRKp during colistin treatment and its rapid evolution within the host.

Molecular phylogeny reveals Varroa mites are not a separate family but a subfamily of Laelapidae.

Varroa mites, notorious for parasitizing honeybees, are generally classified as Varroidae. Their extremely modified morphologies and behaviors have led to debates regarding their phylogenetic position and classification as an independent family. In this study, two different datasets were employed to reconstruct the phylogenies of Varroa mites and related Laelapidae species: (1) 9257 bp from the whole 13 mitochondrial protein-coding genes of 24 taxa, (2) 3158 bp from 113 taxa using Sanger sequencing of four nuclear loci. Both mitochondrial and nuclear analyses consistently place Varroa mites within the Laelapidae. Here we propose to place Varroa mites in the subfamily Varroinae stat. nov., which represents a highly morphologically adapted group within the Laelapidae. Ancestral state reconstructions reveal that bee-associated lifestyles evolved independently at least three times within Laelapidae, with most phoretic traits originating from free-living ancestors. Our revised classification and evolutionary analyses will provide new insight into understanding the Varroa mites.

Higher HIV-1 evolutionary rate is associated with cytotoxic T lymphocyte escape mutations in infants.

Escape from cytotoxic T lymphocyte (CTL) responses toward HIV-1 Gag and Nef has been associated with reduced control of HIV-1 replication in adults. However, less is known about CTL-driven immune selection in infants as longitudinal studies of infants are limited. Here, 1,210 gag and 1,264 nef sequences longitudinally collected within 15 months after birth from 14 HIV-1 perinatally infected infants and their mothers were analyzed. The number of transmitted founder (T/F) viruses and associations between virus evolution, selection, CTL escape, and disease progression were determined. The analyses indicated that a paraphyletic-monophyletic relationship between the mother-infant sequences was common (80%), and that the HIV-1 infection was established by a single T/F virus in 10 of the 12 analyzed infants (83%). Furthermore, most HIV-1 CTL escape mutations among infants were transmitted from the mothers and did not revert during the first year of infection. Still, immune-driven selection was observed at approximately 3 months after HIV-1 infection in infants. Moreover, virus populations with CTL escape mutations in gag evolved faster than those without, independently of disease progression rate. These findings expand the current knowledge of HIV-1 transmission, evolution, and CTL escape in infant HIV-1 infection and are relevant for the development of immune-directed interventions in infants.IMPORTANCEDespite increased coverage in antiretroviral therapy for the prevention of perinatal transmission, paediatric HIV-1 infection remains a significant public health concern, especially in areas of high HIV-1 prevalence. Understanding HIV-1 transmission and the subsequent virus adaptation from the mother to the infant's host environment, as well as the viral factors that affect disease outcome, is important for the development of early immune-directed interventions for infants. This study advances our understanding of vertical HIV-1 transmission, and how infant immune selection pressure is shaping the intra-host evolutionary dynamics of HIV-1.

Secondary messenger signalling influences Pseudomonas aeruginosa adaptation to sinus and lung environments.

Pseudomonas aeruginosa is a cause of chronic respiratory tract infections in people with cystic fibrosis (CF), non-CF bronchiectasis, and chronic obstructive pulmonary disease. Prolonged infection allows the accumulation of mutations and horizontal gene transfer, increasing the likelihood of adaptive phenotypic traits. Adaptation is proposed to arise first in bacterial populations colonizing upper airway environments. Here, we model this process using an experimental evolution approach. Pseudomonas aeruginosa PAO1, which is not airway adapted, was serially passaged, separately, in media chemically reflective of upper or lower airway environments. To explore whether the CF environment selects for unique traits, we separately passaged PAO1 in airway-mimicking media with or without CF-specific factors. Our findings demonstrated that all airway environments-sinus and lungs, under CF and non-CF conditions-selected for loss of twitching motility, increased resistance to multiple antibiotic classes, and a hyper-biofilm phenotype. These traits conferred increased airway colonization potential in an in vivo model. CF-like conditions exerted stronger selective pressures, leading to emergence of more pronounced phenotypes. Loss of twitching was associated with mutations in type IV pili genes. Type IV pili mediate surface attachment, twitching, and induction of cAMP signalling. We additionally identified multiple evolutionary routes to increased biofilm formation involving regulation of cyclic-di-GMP signalling. These included the loss of function mutations in bifA and dipA phosphodiesterase genes and activating mutations in the siaA phosphatase. These data highlight that airway environments select for traits associated with sessile lifestyles and suggest upper airway niches support emergence of phenotypes that promote establishment of lung infection.

Klebsiella pneumoniae exhibiting a phenotypic hyper-splitting phenomenon including the formation of small colony variants.

In this study, we characterized a Klebsiella pneumoniae strain in a patient with shrapnel hip injury, which resulted in multiple phenotypic changes, including the formation of a small colony variant (SCV) phenotype. Although already described since the 1960s, there is little knowledge about SCV phenotypes in Enterobacteriaceae. The formation of SCVs has been recognized as a bacterial strategy to evade host immune responses and compromise the efficacy of antimicrobial therapies, leading to persistent and recurrent courses of infections. In this case, 14 isolates with different resisto- and morpho-types were distinguished from the patient's urine and tissue samples. Whole genome sequencing revealed that all isolates were clonally identical belonging to the K. pneumoniae high-risk sequence type 147. Subculturing the SCV colonies consistently resulted in the reappearance of the initial SCV phenotype and three stable normal-sized phenotypes with distinct morphological characteristics. Additionally, an increase in resistance was observed over time in isolates that shared the same colony appearance. Our findings highlight the complexity of bacterial behavior by revealing a case of phenotypic "hyper-splitting" in a K. pneumoniae SCV and its potential clinical significance.

The effect of molnupiravir and nirmatrelvir on SARS-CoV-2 genome diversity in severe models of COVID-19.

Objectives: Immunocompromised individuals are susceptible to severe COVID-19 and potentially contribute to the emergence of variants with altered pathogenicity due to persistent infection. This study investigated the impact of immunosuppression on SARS-CoV-2 infection in k18-hACE2 mice and the effectiveness of antiviral treatments in this context during the first 7 days of infection. Methods: Mice were immunosuppressed using cyclophosphamide and infected with a B lineage of SARS-CoV-2. Molnupiravir and nirmatrelvir, alone and in combination, were administered and viral load and viral sequence diversity was assessed. Results: Treatment of infected but immune compromised mice with both compounds either singly or in combination resulted in decreased viral loads and pathological changes compared to untreated animals. Treatment also abrogated infection of neuronal tissue. However, no consistent changes in the viral consensus sequence were observed, except for the emergence of the S:H655Y mutation. Molnupiravir, but not nirmatrelvir or immunosuppression alone, increased the transition/transversion (Ts/Tv) ratio, representative of A>G and C>U mutations and this increase was not altered by the co-administration of nirmatrelvir with molnupiravir.Notably, immunosuppression itself did not appear to promote the emergence of mutational characteristic of variants of concern (VOCs). Conclusions: Further investigations are warranted to fully understand the role of immunocompromised individuals in VOC development, especially by taking persistence into consideration, and to inform optimised public health strategies. It is more likely that immunodeficiency promotes viral persistence but does not necessarily lead to substantial consensus-level changes in the absence of antiviral selection pressure. Consistent with mechanisms of action, molnupiravir showed a stronger mutagenic effect than nirmatrelvir in this model.

VIPERA: Viral Intra-Patient Evolution Reporting and Analysis.

Viral mutations within patients nurture the adaptive potential of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) during chronic infections, which are a potential source of variants of concern. However, there is no integrated framework for the evolutionary analysis of intra-patient SARS-CoV-2 serial samples. Herein, we describe Viral Intra-Patient Evolution Reporting and Analysis (VIPERA), a new software that integrates the evaluation of the intra-patient ancestry of SARS-CoV-2 sequences with the analysis of evolutionary trajectories of serial sequences from the same viral infection. We have validated it using positive and negative control datasets and have successfully applied it to a new case, which revealed population dynamics and evidence of adaptive evolution. VIPERA is available under a free software license at https://github.com/PathoGenOmics-Lab/VIPERA.

Dynamic evolution of the sofosbuvir-associated variant A1343V in HEV-infected patients under concomitant sofosbuvir-ribavirin treatment.

Background & Aims: In the absence of a hepatitis E virus (HEV)-specific antiviral treatment, sofosbuvir has recently been shown to have antiviral activity against HEV in vivo. However, a variant, A1343V, that is strongly associated with viral relapse impedes treatment success. In this study, we investigated the occurrence of variants during sofosbuvir and ribavirin treatment in vivo and assessed the sensitivity of resistance-associated variants to concurrent treatment in cell culture. Methods: Two patients with chronic HEV infection that did not clear infection under ribavirin treatment were subsequently treated with a combination of sofosbuvir and ribavirin. We determined response to treatment by measuring liver enzymes and viral load in blood and stool. Moreover, we analyzed viral evolution using polymerase-targeted high-throughput sequencing and assessed replication fitness of resistance-associated variants using a HEV replicon system. Results: Combination treatment was successful in decreasing viral load towards the limit of quantification. However, during treatment sustained virological response was not achieved. Variants associated with sofosbuvir or ribavirin treatment emerged during treatment, including A1343V and G1634R. Moreover, A1343V, as a single or double mutation with G1634R, was associated with sofosbuvir resistance during concomitant treatment in vitro. Conclusions: These results highlight the importance of variant profiling during antiviral treatment of patients with chronic infection. Understanding how intra-host viral evolution impedes treatment success will help guide the design of next-generation antivirals. Impact and implications: The lack of hepatitis E virus (HEV)-specific antivirals to treat chronic infection remains a serious health burden. Although ribavirin, interferon and sofosbuvir have been reported as anti-HEV drugs, not all patients are eligible for treatment or clear infection, since resistant-associated variants can rapidly emerge. In this study, we analyzed the efficacy of sofosbuvir and ribavirin combination treatment in terms of HEV suppression, the emergence of resistance-associated variants and their ability to escape treatment inhibition in vitro. Our results provide novel insights into evolutionary dynamics of HEV during treatment and thus will help guide the design of next-generation antivirals.

Within-host evolutionary dynamics and tissue compartmentalization during acute SARS-CoV-2 infection.

The global evolution of SARS-CoV-2 depends in part upon the evolutionary dynamics within individual hosts with varying immune histories. To characterize the within-host evolution of acute SARS-CoV-2 infection, we sequenced saliva and nasal samples collected daily from vaccinated and unvaccinated individuals early during infection. We show that longitudinal sampling facilitates high-confidence genetic variant detection and reveals evolutionary dynamics missed by less-frequent sampling strategies. Within-host dynamics in both unvaccinated and vaccinated individuals appeared largely stochastic; however, in rare cases, minor genetic variants emerged to frequencies sufficient for forward transmission. Finally, we detected significant genetic compartmentalization of viral variants between saliva and nasal swab sample sites in many individuals. Altogether, these data provide a high-resolution profile of within-host SARS-CoV-2 evolutionary dynamics.IMPORTANCEWe detail the within-host evolutionary dynamics of SARS-CoV-2 during acute infection in 31 individuals using daily longitudinal sampling. We characterized patterns of mutational accumulation for unvaccinated and vaccinated individuals, and observed that temporal variant dynamics in both groups were largely stochastic. Comparison of paired nasal and saliva samples also revealed significant genetic compartmentalization between tissue environments in multiple individuals. Our results demonstrate how selection, genetic drift, and spatial compartmentalization all play important roles in shaping the within-host evolution of SARS-CoV-2 populations during acute infection.

Within-Host Rhinovirus Evolution in Upper and Lower Respiratory Tract Highlights Capsid Variability and Mutation-Independent Compartmentalization.

BACKGROUND: Rhinovirus (RV) infections can progress from the upper (URT) to lower (LRT) respiratory tract in immunocompromised individuals, causing high rates of fatal pneumonia. Little is known about how RV evolves within hosts during infection. METHODS: We sequenced RV complete genomes from 12 hematopoietic cell transplant patients with infection for up to 190 days from both URT (nasal wash, NW) and LRT (bronchoalveolar lavage, BAL). Metagenomic and amplicon next-generation sequencing were used to track the emergence and evolution of intrahost single nucleotide variants (iSNVs). RESULTS: Identical RV intrahost populations in matched NW and BAL specimens indicated no genetic adaptation is required for RV to progress from URT to LRT. Coding iSNVs were 2.3-fold more prevalent in capsid over nonstructural genes. iSNVs modeled were significantly more likely to be found in capsid surface residues, but were not preferentially located in known RV-neutralizing antibody epitopes. Newly emergent, genotype-matched iSNV haplotypes from immunocompromised individuals in 2008-2010 could be detected in Seattle-area community RV sequences in 2020-2021. CONCLUSIONS: RV infections in immunocompromised hosts can progress from URT to LRT with no specific evolutionary requirement. Capsid proteins carry the highest variability and emergent mutations can be detected in other, including future, RV sequences.

Within-Host Resistance and Virulence Evolution of a Hypervirulent Carbapenem-Resistant Klebsiella pneumoniae ST11 Under Antibiotic Pressure.

Background: Hypervirulent carbapenem-resistant Klebsiella pneumoniae (hv-CRKP) has recently aroused an extremely severe health challenge and public concern. However, the underlying mechanisms of fitness costs that accompany antibiotic resistance acquisition remain largely unexplored. Here, we report a hv-CRKP-associated fatal infection and reveal a reduction in virulence due to the acquisition of aminoglycoside resistance. Methods: The bacterial identification, antimicrobial susceptibility, hypermucoviscosity, virulence factors, MLST and serotypes were profiled.The clonal homology and plasmid acquisition among hv-CRKP strains were detected by XbaI and S1-PFGE. The virulence potential of the strains was evaluated using Galleria mellonella larvae infection model, serum resistance assay, capsular polysaccharide quantification, and biofilm formation assay. Genomic variations were identified using whole-genome sequencing (WGS). Results: Four K. pneumoniae carbapenemase (KPC)-producing CRKP strains were consecutively isolated from an 86-year-old patient with severe pneumonia. Whole-genome sequencing (WGS) showed that all four hv-CRKP strains belonged to the ST11-KL64 clone. PFGE analysis revealed that the four ST11-KL64 hv-CRKP strains could be grouped into the same PFGE type. Under the pressure of antibiotics, the antimicrobial resistance of the strains increased and the virulence potential decreased. Further sequencing, using the Nanopore platform, was performed on three representative isolates (WYKP586, WYKP589, and WYKP594). Genomic analysis showed that the plasmids of these three strains underwent a large number of breaks and recombination events under antibiotic pressure. We found that as aminoglycoside resistance emerged via acquisition of the rmtB gene, the hypermucoviscosity and virulence of the strains decreased because of internal mutations in the rmpA and rmpA2 genes. Conclusion: This study shows that ST11-KL64 hv-CRKP can further evolve to acquire aminoglycoside resistance accompanied by decreased virulence to adapt to antibiotic pressure in the host.

Within-host acquisition of colistin-resistance of an NDM-producing Klebsiella quasipneumoniae subsp. similipneumoniae strain through the insertion sequence-903B-mediated inactivation of mgrB gene in a lung transplant child in China.

Background: Colistin, as the antibiotic of "last resort" for carbapenem-resistant Klebsiella, develop resistance during administration of this antimicrobial agent. We identified an NDM-1-producing Klebsiella quasipneumonuae subsp. similipneumoniae (KQSS) strain KQ20605 recovered from a child, which developed resistance to colistin (KQ20786) through acquiring an IS903B element between the -27th and -26th bp of mgrB promoter region after 6-day colistin usage. Objectives: The aim of this study is to explore the source of IS903B in the disruptive mgrB gene and its underlying mechanisms. Materials and methods: Antibiotics susceptibility testing was conducted via microbroth dilution method. The in vitro colistin-induced experiment of KQ20605 was performed to mimic the in vivo transition from colistin-sensitive to resistant. Whole-genome sequencing was used to molecular identification of colistin resistance mechanism. Results: The IS903B element integrated into mgrB gene of KQ20786 had a 100% nucleotide identity and coverage match with one IS903B on plasmid IncR, and only 95.1% (1005/1057) identity to those on chromosome. In vitro, upon the pressure of colistin, KQ20605 could also switch its phenotype from colistin-sensitive to resistant with IS elements (e.g., IS903B and IS26) frequently inserted into mgrB gene at "hotspots", with the insertion site of IS903B nearly identical to that of KQ20786. Furthermore, IS26 elements in this isolate were only encoded by plasmids, including IncR and conjugative plasmid IncN harboring bla NDM. Conclusion: Mobilizable IS elements on plasmids tend to be activated and integrated into mgrB gene at "hotspots" in this KQSS, thereby causing the colistin resistance emergence and further dissemination.

Investigation of the individual genetic evolution of SARS-CoV-2 in a small cluster during the rapid spread of the BF.5 lineage in Tokyo, Japan.

There has been a decreasing trend in new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) cases and fatalities worldwide. The virus has been evolving, indicating the potential emergence of new variants and uncertainties. These challenges necessitate continued efforts in disease control and mitigation strategies. We investigated a small cluster of SARS-CoV-2 Omicron variant infections containing a common set of genomic mutations, which provided a valuable model for investigating the transmission mechanism of genetic alterations. We conducted a study at a medical center in Japan during the Omicron surge (sub-lineage BA.5), sequencing the entire SARS-CoV-2 genomes from infected individuals and evaluating the phylogenetic tree and haplotype network among the variants. We compared the mutations present in each strain within the BA.5 strain, TKYnat2317, which was first identified in Tokyo, Japan. From June 29th to July 4th 2022, nine healthcare workers (HCWs) tested positive for SARS-CoV-2 by real-time PCR. During the same period, five patients also tested positive by real-time PCR. Whole genome sequencing revealed that the infected patients belonged to either the isolated BA.2 or BA.5 sub-lineage, while the healthcare worker infections were classified as BF.5. The phylogenetic tree and haplotype network clearly showed the specificity and similarity of the HCW cluster. We identified 12 common mutations in the cluster, including I110V in nonstructural protein 4 (nsp4), A1020S in the Spike protein, and H47Y in ORF7a, compared to the BA.5 reference. Additionally, one case had the extra nucleotide-deletion mutation I27* in ORF10, and low frequencies of genetic alterations were also found in certain instances. The results of genome sequencing showed that the nine HCWs shared a set of genetic mutations, indicating transmission within the cluster. Minor mutations observed in five HCW individuals suggested the emergence of new virus variants. Five amino acid substitutions occurred in nsp3, which could potentially affect virus replication or immune escape. Intra-host evolution also generated additional mutations. The cluster exhibited a mild disease course, with individuals in this case, recovering without requiring any medical treatments. Further investigation is needed to understand the relationship between the genetic evolution of the virus and the symptoms.

Case Commentary: The hidden side of oxacillin resistance in Staphylococcus aureus.

Acquisition of PBP2a (encoded by the mec gene) is the key resistance mechanism to ß-lactams in Staphylococcus aureus. The mec gene can be easily detected by PCR assays; however, these tools will miss mec-independent oxacillin resistance. This phenotype is mediated by mutations in cell wall metabolism genes that can be acquired during persistent infections under prolonged antibiotic exposure. The complex case presented by Hess et al. (Antimicrob Agents Chemother 67:e00437-23, 2023, https://doi.org/10.1128/aac.00437-23) highlights the diagnostic and therapeutic challenges in the management of mec-independent oxacillin resistance.

Within-Host Evolution of SARS-CoV-2 in a B-Cell Depleted Patient With Successful Treatment.

Prolonged viral shedding of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in an immunocompromised host is a challenge as the treatment and infection control for chronic coronavirus disease 2019 infection is not well established and there is a potential risk of new variants emerging. A 48-year-old woman who underwent chemotherapy, including rituximab and steroid, had reactivation of SARS-CoV-2 68 days after the virus was first detected. She successfully recovered after receiving convalescent plasma and intravenous immunoglobulin. Genomic analysis demonstrated that viruses collected from the nasopharyngeal specimens at day 0 and day 68 had 18 different nucleotide mutations, implying within-host evolution after in-depth epidemiologic investigation.

Pulsed antibiotic treatments of gnotobiotic mice manifest in complex bacterial community dynamics and resistance effects.

Bacteria can evolve to withstand a wide range of antibiotics (ABs) by using various resistance mechanisms. How ABs affect the ecology of the gut microbiome is still poorly understood. We investigated strain-specific responses and evolution during repeated AB perturbations by three clinically relevant ABs, using gnotobiotic mice colonized with a synthetic bacterial community (oligo-mouse-microbiota). Over 80 days, we observed resilience effects at the strain and community levels, and we found that they were correlated with modulations of the estimated growth rate and levels of prophage induction as determined from metagenomics data. Moreover, we tracked mutational changes in the bacterial populations, and this uncovered clonal expansion and contraction of haplotypes and selection of putative AB resistance-conferring SNPs. We functionally verified these mutations via reisolation of clones with increased minimum inhibitory concentration (MIC) of ciprofloxacin and tetracycline from evolved communities. This demonstrates that host-associated microbial communities employ various mechanisms to respond to selective pressures that maintain community stability.

Viral intra-host evolutionary dynamics revealed via serial passage of Japanese encephalitis virus in vitro.

Analyses of viral inter- and intra-host mutations could better guide the prevention and control of infectious diseases. For a long time, studies on viral evolution have focused on viral inter-host variations. Next-generation sequencing has accelerated the investigations of viral intra-host diversity. However, the theoretical basis and dynamic characteristics of viral intra-host mutations remain unknown. Here, using serial passages of the SA14-14-2 vaccine strain of Japanese encephalitis virus (JEV) as the in vitro model, the distribution characteristics of 1,788 detected intra-host single-nucleotide variations (iSNVs) and their mutated frequencies from 477 deep-sequenced samples were analyzed. Our results revealed that in adaptive (baby hamster kidney (BHK)) cells, JEV is under a nearly neutral selection pressure, and both non-synonymous and synonymous mutations represent an S-shaped growth trend over time. A higher positive selection pressure was observed in the nonadaptive (C6/36) cells, and logarithmic growth in non-synonymous iSNVs and linear growth in synonymous iSNVs were observed over time. Moreover, the mutation rates of the NS4B protein and the untranslated region (UTR) of the JEV are significantly different between BHK and C6/36 cells, suggesting that viral selection pressure is regulated by different cellular environments. In addition, no significant difference was detected in the distribution of mutated frequencies of iSNVs between BHK and C6/36 cells.

"Resistance Is Futile": Weaker Selection for Resistance by Abundant Parasites Increases Prevalence and Depresses Host Density.

AbstractTheory often predicts that host populations should evolve greater resistance when parasites become abundant. Furthermore, that evolutionary response could ameliorate declines in host populations during epidemics. Here, we argue for an update: when all host genotypes become sufficiently infected, higher parasite abundance can select for lower resistance because its cost exceeds its benefit. We illustrate such a "resistance is futile" outcome with mathematical and empirical approaches. First, we analyzed an eco-evolutionary model of parasites, hosts, and hosts' resources. We determined eco-evolutionary outcomes for prevalence, host density, and resistance (mathematically, "transmission rate") along ecological and trait gradients that alter parasite abundance. With high enough parasite abundance, hosts evolve lower resistance, amplifying infection prevalence and decreasing host density. In support of these results, a higher supply of nutrients drove larger epidemics of survival-reducing fungal parasites in a mesocosm experiment. In two-genotype treatments, zooplankton hosts evolved less resistance under high-nutrient conditions than under low-nutrient conditions. Less resistance, in turn, was associated with higher infection prevalence and lower host density. Finally, in an analysis of naturally occurring epidemics, we found a broad, bimodal distribution of epidemic sizes consistent with the resistance is futile prediction of the eco-evolutionary model. Together, the model and experiment, supplemented by the field pattern, support predictions that drivers of high parasite abundance can lead to the evolution of lower resistance. Hence, under certain conditions, the most fit strategy for individual hosts exacerbates prevalence and depresses host populations.

Acinetobacter baumannii adaptation to the host pH microenvironment is mediated by allelic variation in a single residue of BauA protein.

Acinetobacter baumannii has been listed as one of the most critical pathogens in nosocomial infections; however, the key genes and mechanisms to adapt to the host microenvironment lack in-depth understanding. In this study, a total of 76 isolates (from 8 to 12 isolates per patient, spanning 128 to 188 days) were longitudinally collected from eight patients to investigate the within-host evolution of A. baumannii. A total of 70 within-host mutations were identified, 80% of which were nonsynonymous, indicating the important role of positive selection. Several evolutionary strategies of A. baumannii to increase its potential to adapt to the host microenvironment were identified, including hypermutation and recombination. Six genes were mutated in isolates from two or more patients, including two TonB-dependent receptor genes (bauA and BJAB07104_RS00665). In particular, the siderophore receptor gene bauA was mutated in multiple isolates from four patients with three MLST types, and all mutations were at amino acid 391 in ligand-binding sites. With 391T or 391A, BauA was more strongly bound to siderophores, which promoted the iron-absorption activity of A. baumannii at acidic or neutral pH, respectively. Through the A/T mutation at site 391 of BauA, A. baumannii displayed two reversible phases to adapt to distinct pH microenvironments. In conclusion, we demonstrated the comprehensive within-host evolutionary dynamics of A. baumannii, and discovered a key mutation of BauA site 391 as a genetic switch to adapt to different pH values, which may represent a model in the pathogen evolutionary adaption of the host microenvironment.

Avian H7N9 influenza viruses are evolutionarily constrained by stochastic processes during replication and transmission in mammals.

H7N9 avian influenza viruses (AIVs) have caused over 1,500 documented human infections since emerging in 2013. Although wild-type H7N9 AIVs can be transmitted by respiratory droplets in ferrets, they have not yet caused widespread outbreaks in humans. Previous studies have revealed molecular determinants of H7N9 AIV host switching, but little is known about potential evolutionary constraints on this process. Here, we compare patterns of sequence evolution for H7N9 AIV and mammalian H1N1 viruses during replication and transmission in ferrets. We show that three main factors-purifying selection, stochasticity, and very narrow transmission bottlenecks-combine to severely constrain the ability of H7N9 AIV to effectively adapt to mammalian hosts in isolated, acute spillover events. We find rare evidence of natural selection favoring new, potentially mammal-adapting mutations within ferrets but no evidence of natural selection acting during transmission. We conclude that human-adapted H7N9 viruses are unlikely to emerge during typical spillover infections. Our findings are instead consistent with a model in which the emergence of a human-transmissible virus would be a rare and unpredictable, though highly consequential, 'jackpot' event. Strategies to control the total number of spillover infections will limit opportunities for the virus to win this evolutionary lottery.