Genomic Characterization of Gonococci from Different Anatomic Sites, Italy, 2007-2014.

In recent decades, Neisseria gonorrhoeae has developed resistance to several antimicrobial classes. Molecular epidemiology approaches are useful for detecting emerging, often resistant, gonococcal clones. In this study, 67 N. gonorrhoeae isolates from different anatomic sites, collected over 8 years in Italy, were analyzed by whole genome sequencing (WGS). WGS was performed using the Illumina NextSeq 500 platform. Phylogenetic analysis was based on core single nucleotide polymorphism (SNP) and core genome multilocus sequence typing (cgMLST). N. gonorrhoeae multi-antigen sequence typing (NG-MAST), MLST, and N. gonorrhoeae sequence typing for antimicrobial resistance (NG-STAR) were carried out in silico using WGS data. Antimicrobial susceptibility against a four-drug panel was evaluated using a gradient diffusion method. Overall, gonococci clustered in accordance with NG-MAST, MLST, NG-STAR, and antimicrobials susceptibility profiles, but not with the site of isolation, HIV status, and patient sexual orientation. Phylogenetic analysis identified nine clades: two of them were the predominant and including gonococci of G1407 and G2400 genogroups.

Whole genome sequencing for the molecular characterization of carbapenem-resistant Klebsiella pneumoniae strains isolated at the Italian ASST Fatebenefratelli Sacco Hospital, 2012-2014.

BACKGROUND: The emergence of carbapenem-resistant Klebsiella pneumoniae strains is threatening antimicrobial treatment. METHODS: Sixty-eight carbapenemase-producing K. pneumoniae strains isolated at Luigi Sacco University Hospital-ASST Fatebenefratelli Sacco (Milan, Italy) between 2012 and 2014 were characterised microbiologically and molecularly. They were tested for drug susceptibility and carbapenemase phenotypes, investigated by means of repetitive extra-genic palindromic polymerase chain reaction (REP-PCR), and fully sequenced by means of next-generation sequencing for the in silico analysis of multi-locus sequence typing (MLST), their resistome, virulome and plasmid content, and their core single nucleotide polymorphism (SNP) genotypes. RESULTS: All of the samples were resistant to carbapenems, other ß-lactams and ciprofloxacin; many were resistant to aminoglycosides and tigecycline; and seven were resistant to colistin. Resistome analysis revealed the presence of blaKPC genes and, less frequently blaSHV, blaTEM, blaCTX-M and blaOXA, which are related to resistance to carbapenem and other ß-lactams. Other genes conferring resistance to aminoglycoside, fluoroquinolone, phenicol, sulphonamide, tetracycline, trimethoprim and macrolide-lincosamide-streptogramin were also detected. Genes related to AcrAB-TolC efflux pump-dependent and pump-independent tigecycline resistance mechanisms were investigated, but it was not possible to clearly correlate the genomic features with tigecycline resistance because of the presence of a common mutation in susceptible, intermediate and resistant strains. Concerning colistin resistance, the mgrB gene was disrupted by an IS5-like element, and the mobile mcr-1 and mcr-2 genes were not detected in two cases. The virulome profile revealed type-3 fimbriae and iron uptake system genes, which are important during the colonisation stage in the mammalian host environment. The in silico detected plasmid replicons were classified as IncFIB(pQil), IncFIB(K), ColRNAI, IncX1, IncX3, IncFII(K), IncN, IncL/M(pMU407) and IncFIA(HI1). REP-PCR showed five major clusters, and MLST revealed six different sequence types: 512, 258, 307, 1519, 745 and 101. Core SNP genotyping, which led to four clusters, correlated with the MLST data. Isolates of the same sequencing type often had common genetic traits, but the SNP analysis allowed greater strain tracking and discrimination than either the REP-PCR or MLST analysis. CONCLUSION: Our findings support the importance of implementing bacterial genomics in clinical medicine in order to complement traditional methods and overcome their limited resolution.

Meningococci of Serogroup X Clonal Complex 181 in Refugee Camps, Italy.

Four cases of infection with serogroup X meningococci (MenX) (1 in 2015 and 3 in 2016) occurred in migrants living in refugee camps or reception centers in Italy. All MenX isolates were identified as clonal complex 181. Our report suggests that serogroup X represents an emerging health threat for persons arriving from African countries.

Molecular characterization of a collection of Neisseria meningitidis isolates from Croatia, June 2009 to January 2014.

In the last decade, the incidence of invasive meningococcal disease (IMD) in Croatia remained stable at approximately 1 case per 100 000 inhabitants, affecting mainly children aged ≤5 years. We report the molecular characterization of meningococci causing IMD occurring from June 2009 to January 2014 in Croatia. Genomic DNA from 50 clinical isolates was analysed for serogroup, multilocus sequence typing and allele type of the two outer membrane protein genes, porA and the iron-regulated fetA. Furthermore, 22 of them were characterized by using whole-genome sequencing to define the meningococcal vaccine four-component meningococcal serogroup B (4CMenB) antigen genes factor H-binding protein (fHbp), Neisseria heparin-binding antigen (nhba) and Neisseria adhesin A (nadA) and the antimicrobial target resistance genes for penicillin (penicillin binding protein 2, penA), ciprofloxacin (DNA gyrase subunit A, gyrA) and rifampicin (ß-subunit of RNA polymerase, rpoB). The Etest was used to phenotypically determine the antimicrobial susceptibility of isolated meningococci. The main serogroup/clonal complex combinations were MenB cc41/44, MenC/cc11, MenW/cc174 and MenY/cc23. PorA P1.7-2, FetA F5-5 and F1-5 were the most represented through the serogroups. Meningococci with decreased susceptibility to penicillin (38.9 %) and one strain resistant to ciprofloxacin were identified. Forty-two percent of MenB showed the presence of at least one of the 4CMenB vaccine antigens (fHbp, NHBA, NadA and PorA). Our findings highlight the genetic variability of meningococci causing IMD in Croatia, especially for the serogroup B. Molecular-based characterization of meningococci is crucial to enhance IMD surveillance and to better plan national immunization programmes.

Genomic characterization of Italian Clostridium botulinum group I strains.

Clostridium botulinum is a gram-positive bacterium capable of producing the botulinum neurotoxin, a powerful poison that causes botulism, a severe neuroparalytic disease. Its genome has been sequenced entirely and its gene content has been analyzed. To date, 19 full genomes and 64 draft genomes are available. The geographical origin of these genomes is predominantly from the US. In the present study, 10 Italian genomes of C. botulinum group I were analyzed and compared with previously sequenced group I genomes, in order to genetically characterize the Italian population of C. botulinum group I and to investigate the phylogenetic relationships among different lineages. Using the suites of software ClonalFrame and ClonalOrigin to perform genomic analysis, we demonstrated that Italian C. botulinum group I population is phylogenetically heterogeneous encompassing different and distant lineages including overseas strains, too. Moreover, a high recombination rate was demonstrated in the evolution of C. botulinum group I species. Finally, genome sequencing of the strain 357 led us to identify a novel botulinum neurotoxin subtype, F8.

Draft Genome Sequence of Clostridium botulinum B2 450 Strain from Wound Botulism in a Drug User in Italy.

Here, we report the draft genome sequence of Clostridium botulinum B2 450, responsible for the first reported case of wound botulism in a drug user in Italy.

Whole-Genome Sequence of Clostridium botulinum A2B3 87, a Highly Virulent Strain Involved in a Fatal Case of Foodborne Botulism in Italy.

Here, we report the genome sequence of a rare bivalent strain of Clostridium botulinum, A2B3 87. The strain was isolated from a foodborne botulism case that occurred in Italy in 1995. The case was characterized by rapid evolution of the illness and failure of conventional treatments.