A novel IncA/C1 group conjugative plasmid, encoding VIM-1 metallo-beta-lactamase, mediates the acquisition of carbapenem resistance in ST104 Klebsiella pneumoniae isolates from neonates in the intensive care unit of V. Monaldi Hospital in Naples

The emergence of carbapenemase producing Enterobacteriaceae has raised major public health concern. The aim of this study was to investigate the molecular epidemiology and the mechanism of carbapenem resistance acquisition of multidrug-resistant Klebsiella pneumoniae isolates from 20 neonates in the neonatal intensive care unit (NICU) of the V. Monaldi Hospital in Naples, Italy, from April 2015 to March 2016. Genotype analysis by pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST) identified PFGE type A and subtypes A1 and A2 in 17, 2, and 1 isolates, respectively, and assigned all isolates to sequence type (ST) 104. K. pneumoniae isolates were resistant to all classes of β-lactams including carbapenems, fosfomycin, gentamicin, and trimethoprim-sulfamethoxazole, but susceptible to quinolones, amikacin, and colistin. Conjugation experiments demonstrated that resistance to third-generation cephems and imipenem could be transferred along with an IncA/C plasmid containing the extended spectrum β-lactamase blaSHV -12 and carbapenem-hydrolyzing metallo-β-lactamase blaV IM-1 genes. The plasmid that we called pIncAC_KP4898 was 156,252 bp in size and included a typical IncA/C backbone, which was assigned to ST12 and core genome (cg) ST12.1 using the IncA/C plasmid MLST (PMLST) scheme. pIncAC_KP4898 showed a mosaic structure with blaV IM-1 into a class I integron, blaSHV -12 flanked by IS6 elements, a mercury resistance and a macrolide 2'-phosphotransferase clusters, ant(3″), aph(3″), aacA4, qnrA1, sul1, and dfrA14 conferring resistance to aminoglycosides, quinolones, sulfonamides, and trimethoprim, respectively, several genes predicted to encode transfer functions and proteins involved in DNA transposition. The acquisition of pIncAC_KP4898 carrying blaV IM-1 and blaSHV -12 contributed to the spread of ST104 K. pneumoniae in the NICU of V. Monaldi Hospital in Naples

The COVID-19 Pandemic Sparked off a Large-Scale Outbreak of Carbapenem-Resistant Acinetobacter baumannii from the Endemic Strains at an Italian Hospital

Acinetobacter baumannii is a nosocomial pathogen that poses a serious threat due to the rise of incidence of multidrug-resistant (MDR) strains. During the COVID-19 pandemic, MDR A. baumannii clones have caused several outbreaks worldwide. Here, we describe a detailed investigation of an MDR A. baumannii outbreak that occurred at Policlinico San Matteo (Pavia, Italy). A total of 96 A. baumannii strains, isolated between January and July 2020 from 41 inpatients (both SARS-CoV-2 positive and negative) in different wards, were characterized by phenotypic and genomic analyses combining Illumina and Nanopore sequencing. Antibiotic susceptibility testing revealed that all isolates were resistant to carbapenems, and the sequence analysis attributed this to the carbapenemase gene blaOXA-23. Virulence factor screening unveiled that all strains carried determinants for biofilm formation, while plasmid analysis revealed the presence of two plasmids, one of which was ~100 kbp long and encoded a phage sequence. A core genome-based phylogeny was inferred to integrate outbreak strain genomes with background genomes from public databases and the local surveillance program. All strains belonged to the globally disseminated sequence type 2 (ST2) clone and were mainly divided into two clades. Isolates from the outbreak clustered with surveillance isolates from 2019, suggesting that the outbreak was caused by two strains that were already circulating in the hospital before the start of the pandemic. The intensive spread of A. baumannii in the hospital was enhanced by the extreme emergency situation of the first COVID-19 pandemic wave that resulted in reduced attention to infection prevention and control practices. IMPORTANCE: The COVID-19 pandemic, especially during the first wave, posed a great challenge to the hospital management and generally promoted nosocomial pathogen dissemination. MDR A. baumannii can easily spread and persist for a long time on surfaces, causing outbreaks in health care settings. Infection prevention and control practices, epidemiological surveillance, and microbiological screening are fundamental in order to control such outbreaks. Here, we sequenced the genomes of 96 isolates from an outbreak of MDR A. baumannii strains using both short- and long-read technology in order to reconstruct the outbreak events in fine detail. The sequence data demonstrated that two endemic clones of MDR A. baumannii were the source of this large hospital outbreak during the first COVID-19 pandemic wave, confirming the effect of COVID-19 emergency disrupting the protection provided by the use of the standard prevention procedures

Presence of wolbachia in three hymenopteran species : diprion pini (Hymenoptera: Diprionidae), neodiprion sertifer (Hymenoptera: Diprionidae), and dahlbominus fuscipennis (Hymenoptera: Eulophidae)

Sawflies are important pests of various plant species. Diprion pini (L.) and Neodiprion sertifer (Geoffroy) (Hymenoptera: Diprionidae) are two of the most important sawfly pests in Italy, and both species are parasitized by the hymenopteran parasitoid Dahlbominus fuscipennis (Zetterstedt). Bacterial endosymbionts are currently studied for their high potential in strategies of biocontrol in a number of insect species. In this study, we investigated the presence of symbiotic bacteria (Wolbachia and Cardinium) in the three species of hymenoptera mentioned earlier, both in wild and laboratory populations. Although all samples were negative for the presence of Cardinium, 100% prevalence for Wolbachia was detected, as all examined individuals resulted to be PCR positive. Furthermore, 16S rDNA and ftsZ gene sequencing indicated that all individuals from the three hymenopteran species are infected by a single Wolbachia strain. Additionally, we report the presence of gynandromorphic individuals in D. pini, both in wild and laboratory-reared populations. Heat treatments on D. pini colonies removed the Wolbachia symbionts, but they also prevented the development of adults

Shifts of Faecal Microbiota during Sporadic Colorectal Carcinogenesis

Gut microbiota has been implicated in the etiopathogenesis of colorectal cancer. The development of colorectal cancer is a multistep process by which healthy epithelium slowly develops into preneoplastic lesions, which in turn progress into malignant carcinomas over time. In particular, sporadic colorectal cancers can arise from adenomas (about 85% of cases) or serrated polyps through the "adenoma-carcinoma" or the "serrated polyp-carcinoma" sequences, respectively. In this study, we performed 16 S rRNA gene sequencing of bacterial DNA extracted from faecal samples to compare the microbiota of healthy subjects and patients with different preneoplastic and neoplastic lesions. We identified putative microbial biomarkers associated with stage-specific progression of colorectal cancer. In particular, bacteria belonging to the Firmicutes and Actinobacteria phyla, as well as members of the Lachnospiraceae family, proved to be specific of the faecal microbiota of patients with preneoplastic lesions, including adenomas and hyperplastic polyps. On the other hand, two families of the Proteobacteria phylum, Alcaligeneaceae and Enterobacteriaceae, with Sutterella and Escherichia/Shigella being the most representative genera, appeared to be associated with malignancy. These findings, once confirmed on larger cohorts of patients, can represent an important step towards the development of more effective diagnostic strategies

Comparative analysis of the two Acinetobacter baumannii multilocus sequence typing (MLST) schemes

Acinetobacter species assigned to the Acinetobacter calcoaceticus-baumannii (Acb) complex, are Gram-negative bacteria responsible for a large number of human infections. The population structure of Acb has been studied using two 7-gene MLST schemes, introduced by Bartual and coworkers (Oxford scheme) and by Diancourt and coworkers (Pasteur scheme). The schemes have three genes in common but underlie two coexisting nomenclatures of sequence types and clonal complexes, which complicates communication on A. baumannii genotypes. The aim of this study was to compare the characteristics of the two schemes to make a recommendation about their usage. Using genome sequences of 730 strains of the Acb complex, we evaluated the phylogenetic congruence of MLST schemes, the correspondence between sequence types, their discriminative power and genotyping reliability from genomic sequences. In silico ST re-assignments highlighted the presence of a second copy of the Oxford gdhB locus, present in 553/730 genomes that has led to the creation of artefactual profiles and STs. The reliability of the two MLST schemes was tested statistically comparing MLST-based phylogenies to two reference phylogenies (core-genome genes and genome-wide SNPs) using topology-based and likelihood-based tests. Additionally, each MLST gene fragment was evaluated by correlating the pairwise nucleotide distances between each pair of genomes calculated on the core-genome and on each single gene fragment. The Pasteur scheme appears to be less discriminant among closely related isolates, but less affected by homologous recombination and more appropriate for precise strain classification in clonal groups, which within this scheme are more often correctly monophyletic. Statistical tests evaluate the tree deriving from the Oxford scheme as more similar to the reference genome trees. Our results, together with previous work, indicate that the Oxford scheme has important issues: gdhB paralogy, recombination, primers sequences, position of the genes on the genome. While there is no complete agreement in all analyses, when considered as a whole the above results indicate that the Pasteur scheme is more appropriate for population biology and epidemiological studies of A. baumannii and related species and we propose that it should be the scheme of choice during the transition toward, and in parallel with, core genome MLST

SteMIDIfactory/P-DOR: P-DOR v1.1

This version is the result of the indications received from reviewers during the publication process of the P-DOR paper

Surveillance in a Neonatal Intensive Care Unit Allowed the Isolation of a Strain of VIM-Producing <i>Pantoea brenneri</i>

Here, we describe the isolation of a strain of the genus Pantoea encoding a VIM carbapenemase, the first to our knowledge. The strain, isolated from a rectal swab of a 10-day-old newborn admitted to a neonatal intensive care unit (NICU), was identified through whole-genome sequencing analyses as Pantoea brenneri. The strain harbored the carbapenemases gene blaVIM-1. The prompt application of contact measures and the isolation of the newborn prevented the dissemination of VIM-producing P. brenneri and of the plasmid carrying the VIM-1 gene to other newborns

Spread of multiple SARS-CoV-2 lineages April-August 2020 anticipated the second pandemic wave in Lombardy (Italy)

During the early phase of the pandemic (20 February-4 April 2020), we have investigated the temporal and geographical evolution of the virus in Lombardy showing the circulation of at least seven lineages distributed differently in the Region. In the present study, the molecular epidemiology of SARS-CoV-2 was monitored in a period between two pandemic waves in order to track the circulation of new variants (April-August 2020). A great majority of SARS-CoV-2 strains (70.8%) belonged to lineages B, B.1, B.1.1 and B.1.1.1, and five strains belonging to four lineages were already reported in Italy (B.1.1.148, B.1.1.162, B.1.1.71, and B.1.425). In addition, 21 SARS-CoV-2 strains belonged to six lineages not previously observed in Italy were detected. No variants of concern were observed. A total of 152/1274 (11.3%) amino acid changes were observed among spike gene sequences and only 26/152 (17.1%) occurred in the receptor-binding domain region of the spike protein. Results of this study are indicative of ongoing transmission throughout the lockdown period, rather than re-introduction of novel lineages past lockdown. The use of molecular epidemiology in Italy should be promoted in order to provide additional understanding of the transmission of the disease and to have major effect on controlling the spread of disease