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STUDY QUESTION How is the semen quality of sexually active men following recovery from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection? SUMMARY ANSWER Twenty-five percent of the men with recent SARS-Cov-2 infections and proven healing were oligo-crypto-azoospermic, despite the absence of virus RNA in semen. WHAT IS KNOWN ALREADY The presence of SARS-CoV-2 in human semen and its role in virus contagion and semen quality after recovery from coronavirus disease 2019 (COVID-19) is still unclear. So far, studies evaluating semen quality and the occurrence of SARS-CoV-2 in semen of infected or proven recovered men are scarce and included a limited number of participants. STUDY DESIGN, SIZE, DURATION A prospective cross-sectional study on 43 sexually active men who were known to have recovered from SARS-CoV2 was performed. Four biological fluid samples, namely saliva, pre-ejaculation urine, semen and post-ejaculation urine, were tested for the SARS-CoV-2 genome. Female partners were retested if any specimen was found to be SARS-CoV-2 positive. Routine semen analysis and quantification of semen leukocytes and interleukin-8 (IL-8) levels were performed. PARTICIPANTS/MATERIALS, SETTING, METHODS Questionnaires including International Index of Erectile Function and Male Sexual Health Questionnaire Short Form were administered to all subjects. The occurrence of virus RNA was evaluated in all the biological fluids collected by RT-PCR. Semen parameters were evaluated according to the World Health Organization manual edition V. Semen IL-8 levels were evaluated by a two-step ELISA method. MAIN RESULTS AND THE ROLE OF CHANCE After recovery from COVID-19, 25% of the men studied were oligo-crypto-azoospermic. Of the 11 men with semen impairment, eight were azoospermic and three were oligospermic. A total of 33 patients (76.7%) showed pathological levels of IL-8 in semen. Oligo-crypto-azoospermia was significantly related to COVID-19 severity (p < 0.001). Three patients (7%) tested positive for at least one sample (one saliva; one pre-ejaculation urine; one semen and one post-ejaculation urine), so the next day new nasopharyngeal swabs were collected. The results from these three patients and their partners were all negative for SARS-CoV-2. LIMITATIONS, REASONS FOR CAUTION Although crypto-azoospermia was found in a high percentage of men who had recovered from COVID-19, clearly exceeding the percentage found in the general population, the previous semen quality of these men was unknown, nor is it known whether a recovery of testicular function was occurring. The low number of enrolled patients may limit the statistical power of study. WIDER IMPLICATIONS OF THE FINDINGS SARS-CoV-2 can be detected in saliva, urine and semen in a small percentage of men who recovered from COVID-19. One-quarter of men who recovered from COVID-19 demonstrated oligo-crypto-azoospermia indicating that an assessment of semen quality should be recommended for men of reproductive age who are affected by COVID-19. STUDY FUNDING/COMPETING INTEREST(S) None TRIAL REGISTRATION NUMBER n/a

A novel resistance gene, named poxtA, encoding a protein of the antibiotic resistance (ARE) ABC-F lineage, was identified in the genome of an MRSA of clinical origin. PoxtA can confer decreased susceptibility to phenicols, oxazolidinones and tetracyclines and is associated with a putative mobile element that could contribute to its horizontal dissemination.

Consecutive non-replicate clinical isolates (n=191) of carbapenem non-susceptible Enterobacteriaceae were collected from 21 hospital laboratories across Italy from November 2013 to April 2014 as part of the European Survey on Carbapenemase-producing Enterobacteriaceae (EuSCAPE) project. Klebsiella pneumoniae carbapenemase-producing K. pneumoniae (KPC-KP) represented 178 (93%) isolates with 76 (43%) respectively resistant to colistin, a key drug for treating carbapenamase-producing Enterobacteriaceae. KPC-KP colistin-resistant isolates were detected in all participating laboratories. This underscores a concerning evolution of colistin resistance in a setting of high KPC-KP endemicity.

A synthetic antimicrobial peptide was identified as a possible candidate for the development of a new antibacterial drug. The peptide, SET-M33L, showed a MIC90 below 1.5 μM and 3 μM for Pseudomonas aeruginosa and Klebsiella pneumoniae, respectively. In in vivo models of P. aeruginosa infections, the peptide and its pegylated form (SET-M33L-PEG) enabled a survival percentage of 60–80% in sepsis and lung infections when injected twice i.v. at 5 mg/Kg, and completely healed skin infections when administered topically. Plasma clearance showed different kinetics for SET-M33L and SET-M33L-PEG, the latter having greater persistence two hours after injection. Bio-distribution in organs did not show significant differences in uptake of the two peptides. Unlike colistin, SET-M33L did not select resistant mutants in bacterial cultures and also proved non genotoxic and to have much lower in vivo toxicity than antimicrobial peptides already used in clinical practice. The characterizations reported here are part of a preclinical development plan that should bring the molecule to clinical trial in the next few years.

Forty-four nonreplicate clinical isolates of Pseudomonas aeruginosa that were resistant to extended-spectrum cephalosporins (ceftazidime and cefepime) and aztreonam, that putatively produced an acquired extended-spectrum ␤-lactamase (ESBL), according to the results of a double-disk synergy test, and that had been involved in nosocomial outbreaks were obtained from six different hospitals in northern Italy and screened for the presence of bla PER ESBL determinants. Twenty isolates, associated with nine independent outbreaks that occurred in five hospitals in the Milan area and its surroundings during 1995-2000, were found to carry an acquired bla PER-1 gene. PER-1 producers representative of the nine outbreaks exhibited a multidrug resistance (MDR) phenotype, including resistance to extended-spectrum cephalosporins, aztreonam, meropenem, aminoglycosides, and in most cases, imipenem and ciprofloxacin. An analysis of macrorestriction profiles of their genomic DNAs by pulsed-field gel electrophoresis revealed an overall clonal diversity of the PER-1 producers, although interhospital clonal spread was also observed. The bla PER-1 gene was not transferable and appeared to be chromosomally located. An analysis of the EcoRI and EcoRV restriction fragment length polymorphisms of the bla PER-1 locus revealed identical patterns for all isolates, and the characterization of a 1.9-kb region containing bla PER-1 revealed a conserved structure in representatives of the various clonal lineages. The present findings indicate that MDR P. aeruginosa clones producing the PER-1 ESBL are endemic to this area of northern Italy, where they have been circulating since the mid-1990s and have been associated with several nosocomial outbreaks.

Acquired metallo-␤-lactamases (MBLs) are resistance determinants of increasing clinical importance in Gram-negative bacterial pathogens, which confer a broad-spectrum ␤-lactam resistance, including carbapenems. Several such enzymes have been described since the 1990s. In the present study, a novel acquired MBL, named FIM-1, was identified and characterized. The bla FIM-1 gene was cloned from a multidrug-resistant Pseudomonas aeruginosa clinical isolate (FI-14/157) cultured from a patient with a vascular graft infection in Florence, Italy. The isolate belonged in the sequence type 235 epidemic clonal lineage. The FIM-1 enzyme is a member of subclass B1 and, among acquired MBLs, exhibited the highest similarity (ca. 40% amino acid identity) with NDM-type enzymes. In P. aeruginosa FI-14/157, the bla FIM-1 gene was apparently inserted into the chromosome and associated with ISCR19-like elements that were likely involved in the capture and mobilization of this MBL gene. Transfer experiments of the bla FIM-1 gene to an Escherichia coli strain or another P. aeruginosa strain by conjugation or electrotransformation were not successful. The FIM-1 protein was produced in E. coli and purified by two chromatography steps. Analysis of the kinetic parameters, carried out with the purified enzyme, revealed that FIM-1 has a broad substrate specificity, with a preference for penicillins (except the 6␣-methoxy derivative temocillin) and carbapenems. Aztreonam was not hydrolyzed. Detection of this novel type of acquired MBL in a P. aeruginosa clinical isolate underscores the increasing diversity of such enzymes that can be encountered in the clinical setting.A cquired metallo-␤-lactamases (MBLs) are resistance determinants of increasing clinical importance in Gram-negative bacterial pathogens, including Pseudomonas aeruginosa, Enterobacteriaceae, and other Gram-negative nonfermenters. These enzymes can degrade most ␤-lactams, including carbapenems, and escape inhibition by the conventional ␤-lactamase inhibitors or avibactam. Thus, they are able to confer an extended ␤-lactam resistance profile to the bacterial host that is not reversible by ␤-lactamase inhibitors (1).Acquired MBLs were detected since the early 1990s, the first representatives being IMP-and VIM-type enzymes (2-4). Thereafter, a number of additional lineages of acquired MBLs have been described, including the SPM-, GIM-, SIM-, KHM-, NDM-, AIM-, DIM-, SMB-, and TMB-type enzymes (5, 6; see also reference 1 and references therein). By convention, different MBL types diverge from each other at by least 30% at the amino acid sequence level (7). Enzymes of different types may differ in functional properties, and the corresponding genes may be associated with different types of mobile genetic elements responsible for their dissemination, such as mobile gene cassettes inserted into integrons, ISCR elements, or composite transposons (8, 9).We report here on the identification and characterization of a new type of acquired MBL, named FIM-1, in a multidrug-resistant clinical ...

Metallo-␤-lactamases (MBLs) can confer resistance to most ␤-lactams, including carbapenems. Their emergence in gram-negative pathogens is a matter of major concern. Italy was the first European country to report the presence of acquired MBLs in gram-negative pathogens and is one of the countries where MBL producers have been detected repeatedly. Here, we present the results of the first Italian nationwide survey of acquired MBLs in gram-negative pathogens. Of 14,812 consecutive nonreplicate clinical isolates (12,245 Enterobacteriaceae isolates and 2,567 gram-negative nonfermenters) screened for reduced carbapenem susceptibility during a 4-month period (September to December 2004), 30 isolates (28 Pseudomonas aeruginosa isolates, 1 Pseudomonas putida isolate, and 1 Enterobacter cloacae isolate) carried acquired MBL determinants. MBL producers were detected in 10 of 12 cities, with a predominance of VIM-type enzymes over IMP-type enzymes (4:1). Although having an overall low prevalence (1.3%) and significant geographical differences, MBL-producing P. aeruginosa strains appeared to be widespread in Italy, with a notable diversity of clones, enzymes, and integrons carrying MBL gene cassettes.␤-Lactamase production is the major mechanism of resistance to ␤-lactam antibiotics in gram-negative pathogens. Carbapenemases, including the metallo-␤-lactamases (MBLs) and the class A and class D serine carbapenemases, are emerging ␤-lactamases of major importance due to their ability to confer resistance to carbapenems in major gram-negative pathogens (27).The acquired MBLs, in particular, are resistance determinants of remarkable clinical concern due to their very broad substrate spectra (including expanded-spectrum cephalosporins and carbapenems) and nonsusceptibility to therapeutic serine-␤-lactamase inhibitors (5,27,30). Acquired MBL genes are usually clustered with other resistance determinants on mobile DNA elements, and their presence is a virtually constant marker for multidrug resistance (27, 30). The first identified acquired MBL, the IMP-1 enzyme, was discovered in the late 1980s in Japan, in multidrug-resistant (MDR) isolates of Pseudomonas aeruginosa (39). Thenceforth, enzymes of the IMP type or of five additional types of acquired MBLs (VIM, SPM, GIM, SIM, and AIM) have been detected worldwide in clinical isolates of gram-negative nonfermenters (GNNFs) showing complex MDR phenotypes and sometimes causing large nosocomial disease outbreaks (27,30,40). The IMP-and VIM-type MBLs are currently the most widespread and have also been detected in several enterobacterial species (30).Although the number of reports on acquired MBLs has steadily increased during the past decade, most reports deal only with sporadic isolates or small outbreaks in individual institutions, while data from larger-scale epidemiological surveys are considerably more limited (5). Italy was the first European country (and the second country after Japan) to report the emergence of acquired MBLs (7,20), as well as one of the countries where MBL-pro...