Rethinking water treatment targets: Bacteria regrowth under unprovable conditions.

Ozonation is among the currently used technologies to remove chemical and biological contaminants from secondary treated urban wastewater (UWW). Despite its effectiveness on the abatement of organic micropollutants (OMPs) and disinfection, previous studies have shown that regrow of bacteria may occur upon storage of the ozonated UWW. This reactivation has been attributed to the high content of assimilable organic carbon after treatment. In order to investigate if ozonation by-products are the main biological regrowth drivers in stored ozonated UWW, the ozonation surviving cells were resuspended in sterile bottled mineral water (MW), simulating a pristine oligotrophic environment. After 7 days storage, organisms such as Acinetobacter, Methylobacterium, Cupriavidus, Massilia, Acidovorax and Pseudomonas were dominant in both ozonated UWW and pristine MW, demonstrating that bacterial regrowth is not strictly related to the eventual presence of ozonation by-products, but instead with the ability of the surviving cells to cope with nutrient-poor environments. The resistome of UWW before and after ozonation was analysed by metagenomic techniques. Draft metagenome assembled genomes (dMAGs), recovered from both ozonated UWW and after cell resuspension in MW, harboured genes conferring resistance to diverse antibiotics classes. Some of these antibiotic resistance genes (ARGs) were located in the vicinity of mobile genetic elements, suggesting their potential to be mobilized. Among these, dMAGs affiliated to taxa with high relative abundance in stored water, such as P. aeruginosa and Acinetobacter spp., harboured ARGs conferring resistance to 12 and 4 families of antibiotics, respectively, including those encoding carbapenem hydrolysing oxacillinases. The results herein obtained point out that the design and development of new wastewater treatment technologies should include measures to attenuate the imbalance of the bacterial communities promoted by storage of the final treated wastewater, even when applying processes with high mineralization rates.

Comparative genome sequence analysis of several species in the genus Tepidimonas and the description of a novel species Tepidimonas charontis sp. nov.

We performed high-quality genome sequencing of eight strains of the species of the genus Tepidimonas and examined the genomes of closely related strains from the databases to understand why Tepidimonas taiwanensis is the only strain of this genus that utilizes glucose and fructose for growth. We found that the assimilation of these hexoses by T. taiwanensis was due to the presence of two transporters that are absent in all other genomes of strains of members of the genus Tepidimonas examined. Some strains lack genes coding for glucokinase, but the Embden-Meyerhof-Parnas pathway appears to be otherwise complete. The pentose phosphate pathway has a complete set of genes, but genes of the Entner-Doudoroff pathway were not identified in the genomes of any of the strains examined. Genome analysis using average nucleotide identity (ANIb), digital DNA-DNA hybridization (dDDH), average amino acid identity (AAI) and phylogenetic analysis of 400 conserved genes was performed to assess the taxonomic classification of the organisms. Two isolates of the genus Tepidimonas from the hot spring at São Pedro do Sul, Portugal, designated SPSP-6T and SPSPC-18 were also examined in this study. These organisms are mixotrophic, have an optimum growth temperature of about 50 ºC, utilize several organic acids and amino acids for growth but do not grow on sugars. Distinctive phenotypic, 16S rRNA gene sequence and genomic characteristics of strains SPSP-6T and SPSPC-18 lead us to propose a novel species based on strain SPSP-6T for which we recommend the name Tepidimonas charontis sp. nov. (=CECT 9683T=LMG 30884T).

Mutation and recombination in pathogen evolution: Relevance, methods and controversies.

Mutation and recombination drive the evolution of most pathogens by generating the genetic variants upon which selection operates. Those variants can, for example, confer resistance to host immune systems and drug therapies or lead to epidemic outbreaks. Given their importance, diverse evolutionary studies have investigated the abundance and consequences of mutation and recombination in pathogen populations. However, some controversies persist regarding the contribution of each evolutionary force to the development of particular phenotypic observations (e.g., drug resistance). In this study, we revise the importance of mutation and recombination in the evolution of pathogens at both intra-host and inter-host levels. We also describe state-of-the-art analytical methodologies to detect and quantify these two evolutionary forces, including biases that are often ignored in evolutionary studies. Finally, we present some of our former studies involving pathogenic taxa where mutation and recombination played crucial roles in the recovery of pathogenic fitness, the generation of interspecific genetic diversity, or the design of centralized vaccines. This review also illustrates several common controversies and pitfalls in the analysis and in the evaluation and interpretation of mutation and recombination outcomes.

Heterogeneous recombination among Hepatitis B virus genotypes.

The rapid evolution of Hepatitis B virus (HBV) through both evolutionary forces, mutation and recombination, allows this virus to generate a large variety of adapted variants at both intra and inter-host levels. It can, for instance, generate drug resistance or the diverse viral genotypes that currently exist in the HBV epidemics. Concerning the latter, it is known that recombination played a major role in the emergence and genetic diversification of novel genotypes. In this regard, the quantification of viral recombination in each genotype can provide relevant information to devise expectations about the evolutionary trends of the epidemic. Here we measured the amount of this evolutionary force by estimating global and local recombination rates in >4700 HBV complete genome sequences corresponding to nine (A to I) HBV genotypes. Counterintuitively, we found that genotype E presents extremely high levels of recombination, followed by genotypes B and C. On the other hand, genotype G presents the lowest level, where recombination is almost negligible. We discuss these findings in the light of known characteristics of these genotypes. Additionally, we present a phylogenetic network to depict the evolutionary history of the studied HBV genotypes. This network clearly classified all genotypes into specific groups and indicated that diverse pairs of genotypes are derived from a common ancestor (i.e., C-I, D-E and, F-H) although still the origin of this virus presented large uncertainty. Altogether we conclude that the amount of observed recombination is heterogeneous among HBV genotypes and that this heterogeneity can influence on the future expansion of the epidemic.