Predictive modeling of antibiotic eradication therapy success for new-onset Pseudomonas aeruginosa pulmonary infections in children with cystic fibrosis.

Chronic Pseudomonas aeruginosa (Pa) lung infections are the leading cause of mortality among cystic fibrosis (CF) patients; therefore, the eradication of new-onset Pa lung infections is an important therapeutic goal that can have long-term health benefits. The use of early antibiotic eradication therapy (AET) has been shown to clear the majority of new-onset Pa infections, and it is hoped that identifying the underlying basis for AET failure will further improve treatment outcomes. Here we generated machine learning models to predict AET outcomes based on pathogen genomic data. We used a nested cross validation design, population structure control, and recursive feature selection to improve model performance and showed that incorporating population structure control was crucial for improving model interpretation and generalizability. Our best model, controlling for population structure and using only 30 recursively selected features, had an area under the curve of 0.87 for a holdout test dataset. The top-ranked features were generally associated with motility, adhesion, and biofilm formation.

Pseudomonas aeruginosa aggregation and Psl expression in sputum is associated with antibiotic eradication failure in children with cystic fibrosis.

We previously demonstrated that P. aeruginosa isolates that persisted in children with cystic fibrosis (CF) despite inhaled tobramycin treatment had increased anti-Psl antibody binding in vitro compared to those successfully eradicated. We aimed to validate these findings by directly visualizing P. aeruginosa in CF sputum. This was a prospective observational study of children with CF with new-onset P. aeruginosa infection who underwent inhaled tobramycin eradication treatment. Using microbial identification passive clarity technique (MiPACT), P. aeruginosa was visualized in sputum samples obtained before treatment and classified as persistent or eradicated based on outcomes. Pre-treatment isolates were also grown as biofilms in vitro. Of 11 patients enrolled, 4 developed persistent infection and 7 eradicated infection. P. aeruginosa biovolume and the number as well as size of P. aeruginosa aggregates were greater in the sputum of those with persistent compared with eradicated infections (p < 0.01). The amount of Psl antibody binding in sputum was also greater overall (p < 0.05) in samples with increased P. aeruginosa biovolume. When visualized in sputum, P. aeruginosa had a greater biovolume, with more expressed Psl, and formed more numerous, larger aggregates in CF children who failed eradication therapy compared to those who successfully cleared their infection.

Evolutionary History and Strength of Selection Determine the Rate of Antibiotic Resistance Adaptation.

Bacterial adaptation to stressful environments often produces evolutionary constraints whereby increases in resistance are associated with reduced fitness in a different environment. The exploitation of this resistance-cost trade-off has been proposed as the basis of rational antimicrobial treatment strategies designed to limit the evolution of drug resistance in bacterial pathogens. Recent theoretical, laboratory, and clinical studies have shown that fluctuating selection can maintain drug efficacy and even restore drug susceptibility, but can also increase the rate of adaptation and promote cross-resistance to other antibiotics. In this paper, we combine mathematical modeling, experimental evolution, and whole-genome sequencing to follow evolutionary trajectories towards ß-lactam resistance under fluctuating selective conditions. Our experimental model system consists of eight populations of Escherichia coli K12 evolving in parallel to a serial dilution protocol designed to dynamically control the strength of selection for resistance. We implemented adaptive ramps with mild and strong selection, resulting in evolved populations with similar levels of resistance, but with different evolutionary dynamics and diverging genotypic profiles. We found that mutations that emerged under strong selection are unstable in the absence of selection, in contrast to resistance mutations previously selected in the mild selection regime that were stably maintained in drug-free environments and positively selected for when antibiotics were reintroduced. Altogether, our population dynamics model and the phenotypic and genomic analysis of the evolved populations show that the rate of resistance adaptation is contingent upon the strength of selection, but also on evolutionary constraints imposed by prior drug exposures.

Spirochetes isolated from arthropods constitute a novel genus Entomospira genus novum within the order Spirochaetales.

Spirochetal bacteria were successfully isolated from mosquitoes (Culex pipiens, Aedes cinereus) in the Czech Republic between 1999 and 2002. Preliminary 16S rRNA phylogenetic sequence analysis showed that these strains differed significantly from other spirochetal genera within the family Spirochaetaceae and suggested a novel bacterial genus in this family. To obtain more comprehensive genomic information of these isolates, we used Illumina MiSeq and Oxford Nanopore technologies to sequence four genomes of these spirochetes (BR151, BR149, BR193, BR208). The overall size of the genomes varied between 1.68 and 1.78 Mb; the GC content ranged from 38.5 to 45.8%. Draft genomes were compared to 36 publicly available genomes encompassing eight genera from the class Spirochaetes. A phylogeny generated from orthologous genes across all taxa and the percentage of conserved proteins (POCP) confirmed the genus status of these novel spirochetes. The genus Entomospira gen. nov. is proposed with BR151 selected as type species of the genus. For this isolate and the closest related isolate, BR149, we propose the species name Entomospira culicis sp. nov. The two other isolates BR208 and BR193 are named Entomospira nematocera sp. nov. (BR208) and Entomospira entomophilus sp. nov. (BR193). Finally, we discuss their interesting phylogenetic positioning.

Evolutionary and structural analyses of SARS-CoV-2 D614G spike protein mutation now documented worldwide.

The COVID-19 pandemic, caused by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), was declared on March 11, 2020 by the World Health Organization. As of the 31st of May, 2020, there have been more than 6 million COVID-19 cases diagnosed worldwide and over 370,000 deaths, according to Johns Hopkins. Thousands of SARS-CoV-2 strains have been sequenced to date, providing a valuable opportunity to investigate the evolution of the virus on a global scale. We performed a phylogenetic analysis of over 1,225 SARS-CoV-2 genomes spanning from late December 2019 to mid-March 2020. We identified a missense mutation, D614G, in the spike protein of SARS-CoV-2, which has emerged as a predominant clade in Europe (954 of 1,449 (66%) sequences) and is spreading worldwide (1,237 of 2,795 (44%) sequences). Molecular dating analysis estimated the emergence of this clade around mid-to-late January (10-25 January) 2020. We also applied structural bioinformatics to assess the potential impact of D614G on the virulence and epidemiology of SARS-CoV-2. In silico analyses on the spike protein structure suggests that the mutation is most likely neutral to protein function as it relates to its interaction with the human ACE2 receptor. The lack of clinical metadata available prevented our investigation of association between viral clade and disease severity phenotype. Future work that can leverage clinical outcome data with both viral and human genomic diversity is needed to monitor the pandemic.

Rejection of the name Borreliella and all proposed species comb. nov. placed therein.

Rejection (nomen rejiciendum) of the name Borreliella and all new combinations therein is being requested on grounds of risk to human health and patient safety (Principle 1, subprinciple 2 and Rule 56a) and violation to aim for stability of names, to avoid useless creation of names (Principle 1, subprinciple 1 and 3) and that names should not be changed without sufficient reason (Principle 9 of the International Code of Nomenclature of Prokaryotes).

Origin of OXA-23 Variant OXA-239 from a Recently Emerged Lineage of Acinetobacter baumannii International Clone V.

Over the last few decades, carbapenemase-producing Acinetobacter baumannii has become a major cause of nosocomial infections all over the world. However, the genome identity of lineages of this species in Latin America has not been studied as much as in developed countries. Here, through a population genomics approach considering the whole genomes of 148 isolates (almost 40 from Mexico and Honduras), we describe the recent emergence of the lineage sequence type 758 (ST758), which belongs to the international clone V and has spread out to Canada, Mexico, Honduras, and Colombia. Notably, this lineage was found to coexist with other A. baumannii lineages in hospitals in Mexico and Honduras. Isolates from this lineage show considerable variation in antibiotic resistance profiles, but most of them are resistant to carbapenems. Moreover, we found a variety of acquired oxacillinase (OXA) families within this lineage and tracked the very recent inception, and subsequent horizontal transmission, of the OXA-239 carbapenemase. This work highlights the urgent need to investigate recently emerged lineages of this species in Latin America and elsewhere, as these might harbor novel antibiotic resistance genes.IMPORTANCEA. baumannii is a major cause of nosocomial infections all over the world. Although many isolates from developed countries have been studied in terms of their genome sequence, isolates from Latin America have been much less studied. In this study, using a population genomics approach considering the whole genomes of 148 isolates, we describe the recent emergence of the lineage ST758 endemic to Latin America and the inception of the OXA-239 carbapenemase. Our study highlights the urgent need to investigate recently emerged lineages of this species in Latin America and elsewhere, as these might harbor novel antibiotic resistance genes.

High Phenotypic and Genotypic Diversity of Enterococcus faecium from Clinical and Commensal Isolates in Third Level Hospital.

Background: The use of antimicrobials and myeloablative chemotherapy regimens has promoted multiresistant microorganisms to emerge as nosocomial pathogens, such as vancomycin-resistant Enterococcus faecium (VREfm). We described a polyclonal outbreak of bloodstream infection caused by Efm in a hemato-oncological ward in Mexico. Our aim was to describe the clonal complex (CC) of the Efm strains isolated in the outbreak in comparison with commensal and environmental isolates. Methodology: Sixty Efm clinical, environmental, and commensal strains were included. We constructed a cladogram and a phylogenetic tree using Vitek and Multilocus sequence typing data, respectively. Results: We reported 20 new sequence types (ST), among which 17/43 clinical isolates belonged to CC17. The predominant ST in the clinical strains were ST757, ST1304, ST412, and ST770. Neither environmental nor commensal isolates belonged to CC17. The phylogeny of our collection shows that the majority of the clinical isolates were different from the environmental and commensal isolates, and only a small group of clinical isolates was closely related with environmental and commensal isolates. The cladogram revealed a similar segregation to that of the phylogeny. Conclusions: We found a high diversity among clinical, environmental, and commensal strains in a group of samples in a single hospital. Highest diversity was found between commensal and environmental isolates.

Phylogenomics Reveals Clear Cases of Misclassification and Genus-Wide Phylogenetic Markers for Acinetobacter.

The Gram-negative Acinetobacter genus has several species of clear medical relevance. Many fully sequenced genomes belonging to the genus have been published in recent years; however, there has not been a recent attempt to infer the evolutionary history of Acinetobacter with that vast amount of information. Here, through a phylogenomic approach, we established the most up-to-date view of the evolutionary relationships within this genus and highlighted several cases of poor classification, especially for the very closely related species within the Acinetobacter calcoaceticus-Acinetobacter baumannii complex (Acb complex). Furthermore, we determined appropriate phylogenetic markers for this genus and showed that concatenation of the top 13 gives a very decent reflection of the evolutionary relationships for the genus Acinetobacter. The intersection between our top markers and previously defined universal markers is very small. In general, our study shows that, although there seems to be hardly any universal markers, bespoke phylogenomic approaches can be used to infer the phylogeny of different bacterial genera. We expect that ad hoc phylogenomic approaches will be the standard in the years to come and will provide enough information to resolve intricate evolutionary relationships like those observed in the Acb complex.

Unexplored Genetic Diversity of Multidrug- and Extremely Drug-Resistant Acinetobacter baumannii Isolates from Tertiary Hospitals in Honduras.

Although Acinetobacter baumannii has become one of the most important nosocomial pathogens worldwide, very little is known about the genetic identity of isolates from less developed countries in Latin America. To alleviate this, we sequenced the genomes of 16 A. baumannii isolates from Honduras. Whole-genome sequencing was conducted on 16 isolates from five Honduran Hospitals. With the sequences of these Honduran isolates and other 42 publically available genomes, a maximum likelihood phylogeny was constructed to establish the relationship between the Honduran isolates and those belonging to the International Clones (ICs). In addition, sequence type (ST) assignation was conducted by the PubMLST, and antibiotic resistance genes were identified using ResFinder. The Honduran isolates are highly diverse and contain new allele combinations under the Bartual multilocus sequence typing scheme. The most common STs were STB447/STP10 and STB758/STP156. Furthermore, none of these isolates belongs to clonal complexes related to the ICs. Antibiotic susceptibility profiles of these isolates showed that they are multidrug resistant (MDR) or extensively drug resistant (XDR). In addition, the Honduran isolates had genes involved in resistance to seven antibiotic families. For instance, several blaOXA alleles were found, including blaOXA-23 and a gene encoding the metallo-beta-lactamase NDM-1. Notably, nine of the Honduran isolates have antibiotic resistance genes to three or more antibiotic families. In summary, in this study, we unveiled an untapped source of genetic diversity of MDR and XDR isolates; notably, these isolates did not belong to the well-known ICs.

Inaccurate Multilocus Sequence Typing of Acinetobacter baumannii.

Multilocus sequence typing has been useful for genotyping pathogens in surveillance and epidemiologic studies. However, it cannot reflect the true relationships of isolates for species with very dynamic genomes. Using a robust genome phylogeny, we demonstrated the limitations of this method for typing Acinetobacter baumannii.

Phylogenomics picks out the par excellence markers for species phylogeny in the genus Staphylococcus.

Although genome sequencing has become a very promising approach to conduct microbial taxonomy, few labs have the resources to afford this especially when dealing with data sets of hundreds to thousands of isolates. The goal of this study was to identify the most adequate loci for inferring the phylogeny of the species within the genus Staphylococcus; with the idea that those who cannot afford whole genome sequencing can use these loci to carry out species assignation confidently. We retrieved 177 orthologous groups (OGs) by using a genome-based phylogeny and an average nucleotide identity analysis. The top 26 OGs showed topologies similar to the species tree and the concatenation of them yielded a topology almost identical to that of the species tree. Furthermore, a phylogeny of just the top seven OGs could be used for species assignment. We sequenced four staphylococcus isolates to test the 26 OGs and found that these OGs were far superior to commonly used markers for this genus. On the whole, our procedure allowed identification of the most adequate markers for inferring the phylogeny within the genus Staphylococcus. We anticipate that this approach will be employed for the identification of the most suitable markers for other bacterial genera and can be very helpful to sort out poorly classified genera.

Rapid Gene Turnover as a Significant Source of Genetic Variation in a Recently Seeded Population of a Healthcare-Associated Pathogen.

Genome sequencing has been useful to gain an understanding of bacterial evolution. It has been used for studying the phylogeography and/or the impact of mutation and recombination on bacterial populations. However, it has rarely been used to study gene turnover at microevolutionary scales. Here, we sequenced Mexican strains of the human pathogen Acinetobacter baumannii sampled from the same locale over a 3 year period to obtain insights into the microevolutionary dynamics of gene content variability. We found that the Mexican A. baumannii population was recently founded and has been emerging due to a rapid clonal expansion. Furthermore, we noticed that on average the Mexican strains differed from each other by over 300 genes and, notably, this gene content variation has accrued more frequently and faster than the accumulation of mutations. Moreover, due to its rapid pace, gene content variation reflects the phylogeny only at very short periods of time. Additionally, we found that the external branches of the phylogeny had almost 100 more genes than the internal branches. All in all, these results show that rapid gene turnover has been of paramount importance in producing genetic variation within this population and demonstrate the utility of genome sequencing to study alternative forms of genetic variation.

Complete Genome Sequence of a Multidrug-Resistant Acinetobacter baumannii Isolate Obtained from a Mexican Hospital (Sequence Type 422).

Acinetobacter baumannii has emerged as a dangerous nosocomial pathogen, particularly for severely ill patients in intensive care units and patients with hematologic malignancies. Here, we present the complete genome sequence of a multidrug-resistant A. baumannii isolate, recovered from a Mexican hospital and classified as sequence type 422 according to the multilocus sequence typing Pasteur scheme.

First Genome Sequence of a Mexican Multidrug-Resistant Acinetobacter baumannii Isolate.

Acinetobacter baumanniihas emerged as an important nosocomial pathogen worldwide. Here, we present the draft genome of the first multidrug-resistantA. baumanniiisolate, sampled from a tertiary hospital in Mexico City. This genome will provide a starting point for studying the genomic diversity of this species in Mexico.