AUTHOR=Cafiso Viviana , Stracquadanio Stefano , Lo Verde Flavia , Dovere Veronica , Zega Alessandra , Pigola Giuseppe , Aranda Jesús , Stefani Stefania 

TITLE=COLRAcinetobacter baumannii sRNA Signatures: Computational Comparative Identification and Biological Targets

JOURNAL=Frontiers in Microbiology

VOLUME=10

YEAR=2020

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2019.03075

DOI=10.3389/fmicb.2019.03075

ISSN=1664-302X

ABSTRACT=<p>Multidrug-Resistant (MDR) and Extensively Drug Resistant (XDR) <italic>Acinetobacter baumannii</italic> (<italic>Ab</italic>) represent a serious cause of healthcare-associated infections worldwide. Currently, the available treatment options are very restricted and colistin-based therapies are last-line treatments of these infections, even though colistin resistant (COL<sup>R</sup>) <italic>Ab</italic> have rarely been isolated yet. In bacteria, small non-coding RNAs (sRNAs) have been implicated in regulatory pathways of different biological functions, however, no knowledge exists about the sRNA role on the biological adaptation in COL<sup>R</sup><italic>Ab</italic>. Our study investigated two Italian XDR isogenic colistin-susceptible/resistant (COL<sup>S/R</sup>) <italic>Ab</italic> strain-pairs to discover new sRNA signatures. Comparative sRNA transcriptome (sRNAome) analyses were carried out by Illumina RNA-seq using both a Tru-Seq and a Short Insert library, whilst <italic>Ab</italic> ATCC 17978 and ACICU Reference Genome assembly, mapping, annotation and statistically significant differential expression (<italic>q</italic>-value ≤ 0.01) of the raw reads were performed by the Rockhopper tool. A computational filtering, sorting only similarly statistically significant differentially expressed (DE) sRNAs mapping on the same gene in both COL<sup>R</sup><italic>Ab</italic> isolates was conducted. COL<sup>R</sup> vs. COL<sup>S</sup> sRNAome, analyzed integrating the DE sRNAs obtained from the two different libraries, revealed some statistically significant DE sRNAs in COL<sup>R</sup><italic>Ab</italic>. In detail, we found: (i) two different under-expressed <italic>cis</italic>-acting sRNAs (<italic>Ab</italic>sRNA<sub>1</sub> and <italic>Ab</italic>sRNA<sub>2</sub>) mapping in antisense orientation the 16S rRNA gene A1S_r01, (ii) one under-expressed <italic>cis</italic>-acting sRNA (<italic>Ab</italic>sRNA<sub>3</sub>) targeting the A1S_2505 gene (hypothetical protein), (iii) one under-expressed microRNA-size small RNA fragment (<italic>Ab</italic>sRNA<sub>4</sub>) and its pre-micro<italic>Ab</italic>sRNA<sub>4</sub> targeting the A1S_0501 gene (hypothetical protein), (iv) as well as an over-expressed microRNA-size small RNA fragment (<italic>Ab</italic>sRNA<sub>5</sub>) and its pre-micro<italic>Ab</italic>sRNA<sub>5</sub> targeting the A1S_3097 gene (signal peptide). Custom TaqMan<sup>®</sup> probe-based real-time qPCRs validated the expression pattern of the selected sRNA candidates shown by RNA-seq. Furthermore, analysis on sRNA ΔA1S_r01, ΔA1S_2505 as well as the over-expressed A1S_3097 mutants revealed no effects on colistin resistance. Our study, for the first time, found the sRNAome signatures of clinical COL<sup>R</sup><italic>Ab</italic> with a computational prediction of their targets related to protein synthesis, host-microbe interaction and other different biological functions, including biofilm production, cell-cycle control, virulence, and antibiotic-resistance.</p>