One-year surveillance for hypervirulent Klebsiella pneumoniae detected carbapenem-resistant superbugs.

Hypervirulent Klebsiella pneumoniae (hvKp) can cause infections in clinically healthy people, such as young and immunocompetent patients. Genes involved in the capsule synthesis or those encoding the siderophores have been adopted as predictors of hvKp. Certain sequence types, such as ST23 and ST86, have been associated with hvKp strains, too. The aim of this study was to investigate the presence of hvKp among 354 K. pneumoniae strains isolated from clinical samples of patients admitted to an Italian 900-bed hospital between 21 May 2021 and April 2022. All the isolates were screened by PCR for the amplification of virulence loci. Whole genome sequencing was performed in strains tested positive for at least one target gene. Thirteen out of 354 (3.7%) were hvKp. Five were wild type and belonged to the hypervirulent clones ST23, ST86, ST5, and ST375 and to the new clone ST6310. Six strains carried the blaKPC gene: three belonged to ST101, two to ST512, and one to ST395. Two isolates were ST147 and carried the blaNDM gene. Although hvKp isolation is not frequent, their presence should be systematically investigated to avoid the spreading of both virulent strains and strains with combined increase in virulence and resistance to antibiotics. PCR-based protocols are essential for surveillance of these strains, which do not always show a recognizable phenotype. Moreover, hvKp strains were isolated also from patients without history of recent foreign travels, indicating an increased spreading of these strains as well as an underestimated of their circulation so far.IMPORTANCEKlebsiella pneumoniae is a healthcare-associated pathogen frequently resistant to antibiotics. Hypervirulent strains of pneumoniae (hvKp) can spread from the primary site of infection to multiple sites causing life-threatening infections also in young otherwise healthy individuals. This study described the isolation of 13 isolates of K. pneumoniae with increased virulence in a large tertiary hospital over a 1-year period. Among them, eight strains were multidrug resistant and hypervirulent. Although these hypervirulent strains are still rare in Italy, their presence is particularly concerning since they can cause difficult-to-treat life-threatening infections. Moreover, not all the hypervirulent isolates were positive by the string test, so hvKp isolates were not always phenotypically detectable. Molecular biology techniques such as PCR amplification and next generation sequencing are therefore necessary for the detection of hvKp isolates, and surveillance programs exploiting molecular techniques are highly desirable.

Combining Genome Surveillance and Metadata To Characterize the Diversity of Staphylococcus aureus Circulating in an Italian Hospital over a 9-Year Period.

Staphylococcus aureus is an opportunistic pathogen and a leading cause of morbidity and mortality worldwide. Genomic-based surveillance has greatly improved our ability to track the emergence and spread of high-risk clones, but the full potential of genomic data is only reached when used in conjunction with detailed metadata. Here, we demonstrate the utility of an integrated approach by leveraging a curated collection of clinical and epidemiological metadata of S. aureus in the San Matteo Hospital (Italy) through a semisupervised clustering strategy. We sequenced 226 sepsis S. aureus samples, recovered over a period of 9 years. By using existing antibiotic profiling data, we selected strains that capture the full diversity of the population. Genome analysis revealed 49 sequence types, 16 of which are novel. Comparative genomic analyses of hospital- and community-acquired infection ruled out the existence of genomic features differentiating them, while evolutionary analyses of genes and traits of interest highlighted different dynamics of acquisition and loss between antibiotic resistance and virulence genes. Finally, highly resistant clones belonging to clonal complexes (CC) 8 and 22 were found to be responsible for abundant infections and deaths, while the highly virulent CC30 was responsible for rare but deadly episodes of infections. IMPORTANCE Genome sequencing is an important tool in clinical microbiology, as it allows in-depth characterization of isolates of interest and can propel genome-based surveillance studies. Such studies can benefit from ad hoc methods of sample selection to capture the genomic diversity present in a data set. Here, we present an approach based on clustering of antibiotic resistance profiles that allows optimal sample selection for bacterial genomic surveillance. We apply the method to a 9-year collection of Staphylococcus aureus from a large hospital in northern Italy. Our method allows us to sequence the genomes of a large variety of strains of this important pathogen, which we then leverage to characterize the epidemiology in the hospital and to perform evolutionary analyses on genes and traits of interest. These analyses highlight different dynamics of acquisition and loss between antibiotic resistance and virulence genes.

Livestock-associated methicillin-resistant Staphylococcus aureus in inpatients: a snapshot from an Italian hospital.

OBJECTIVES: This study aimed to characterize livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) strains isolated from patients admitted to Policlinico San Matteo in Pavia, located in an Italian region with high livestock density. METHODS: The starting dataset was composed by 353 MRSA strains isolated from blood cultures between 2011 and 2019 and in 954 MRSA isolated from nasal swabs, wound swabs, skin swabs, ulcer swabs, conjunctival swabs, urine and respiratory samples collected between 2018 and 2019. LA-MRSA was identified based on being MRSA resistant to tetracycline and negative for the PCR amplification of scn locus. Whole genome sequencing of the selected strains was performed, and virulence and resistance genes searched. RESULTS: Five out of 353 MRSA isolates from blood cultures (1.4%) and nine out of 904 MRSA isolates obtained from other materials (1%) were resistant to tetracycline and negative for the scn locus. The 14 strains were also negative for the lukS-pv, tsst, eta and etb loci. Nine of the 14 strains belonged to ST398, the most common ST of LA-MRSA in Europe. ST398 isolates belonged to four spa-types, of which the prevalent was t899. Eight genomes had the cassette SCCmec type V, five genomes had SCCmec type IV and one genome lacked SCCmec, mecA and mecC. CONCLUSION: The frequency of LA-MRSA in the patients of this study (1.4% in blood cultures, 1% in other samples) is low but relatively constant over time prevalence and comparable to that found in the few studies performed on patients to date.

Epidemiology and microbiology of prosthetic joint infections: a nine-year, single-center experience in Pavia, Northern Italy.

BACKGROUND: Prosthetic joint infections (PJIs) are a growing matter of concern due to their economic and social burden on health systems. In Italy, surgical data on PJIs are available in a national registry, but microbiological data are still scarce. MATERIALS AND METHODS: We performed a retrospective study at a single center with records of patients treated for primary PJIs of knee or hip from January 1, 2011, to May 30, 2018. Patients with infections of osteosynthesis means and external devices were excluded, as well as PJI recurrences and polytrauma patients. Infections were diagnosed according to IDSA and MSIS criteria. We collected data on demographics, risk factors and microbiology. All patients seen at our center undergo blood cultures and synovial fluid cultures, periarticular biopsy and prosthesis sonication by Bactosonic®. This was used only after 2014. Bacterial identification is achieved by MALDI-TOF, PHOENIX 100 and standard methods. Chi-square or Fisher tests were used to test statistical differences in proportions. RESULTS: Fifty-one patients matched our inclusion criteria. Of these, 16 (31.4%) were enrolled before 2014. The median age was 68.5 (range 22-88). The most common risk factors were obesity (34%), diabetes (21%) and chronic kidney disease (14%). Seventeen patients were diagnosed with a culture-negative PJIs (33.3%). Staphylococcus aureus was the most commonly isolated pathogen (14/51, 27.5%), followed by coagulase-negative staphylococci (7/51, 13.7%). Methicillin-resistant S. aureus rate was 28.6%. The rate of culture-negative PJIs dropped from 56 to 22% after 2014, with a significant difference between the two time periods (p = 0.016). CONCLUSIONS: The introduction of sonication dramatically increased our diagnostic accuracy. Our microbiological data are in line with those from other studies conducted in Italy.

Screening program for latent tuberculosis infection in asylum seekers - a single center experience in Pavia, Italy.

BACKGROUND: The management of Latent Tuberculosis Infection is crucial in fighting Tuberculosis worldwide, and particularly in low incidence European Countries. While guidelines for the management of Tuberculosis in newly arrived immigrants have been issued by the European Center for Disease Control and Prevention and by the National Health Authorities in Italy, these are not widely implemented yet at local level. STUDY DESIGN: We report our program for the screening of Latent Tuberculosis Infection and active Tuberculosis in asylum seekers, jointly implemented by Public Health Authorities and the Infectious Diseases Department of a tertiary care, teaching hospital in Northern Italy. METHODS: We reviewed records of the asylum seekers who were screened at our center via Tuberculin Skin Test and/or Interferon Gamma Release Assay plus chest X-ray and either treated with Isoniazid Preventive Treatment or for active Tuberculosis Disease in case of positive results. RESULTS: We screened 726 migrants, mostly males (97.3%) and from Sub-Saharan Africa (82.2%) and found a high adherence rate for both screening (98.2%) and Isoniazid Preventive Treatment (90.1%). In addition, we found seven cases of active Tuberculosis. CONCLUSIONS: Latent Tuberculosis Infection screening and treatment proved feasible in our program, which should be systematically implemented in asylum seekers reaching Europe.

Prosthetic Joint Infection from Carbapenemase-Resistant Klebsiella pneumoniae Successfully Treated with Ceftazidime-Avibactam.

Antimicrobial resistance in Gram-negative bacteria, particularly Enterobacteriaceae, has become a leading cause of morbidity and mortality and a serious public health concern. Gram-negative bacteria carrying extended-spectrum beta-lactamase (ESBL) enzymes now represent a significant proportion of all bacteria isolated from different countries worldwide. Furthermore, the increasing number of isolates carrying carbapenemases in recent years includes multidrug-resistant (MDR), extensively drug-resistant (XDR), and pandrug-resistant (PDR) bacteria. Here, we describe what, to our knowledge, is the first case of a patient with a prosthetic joint infection from carbapenemase-resistant Klebsiella pneumoniae (CRKP) successfully treated with ceftazidime-avibactam in Italy.

Evaluation of a model to improve collection of blood cultures in patients with sepsis in the emergency room.

Sepsis begins outside of the hospital for nearly 80% of patients and the emergency room (ER) represents the first contact with the health care system. This study evaluates a project to improve collection of blood cultures (BCs) in patients with sepsis in the ER consisting of staff education and completion of the appropriate BC pre-analytical phase. A retrospective observational study performed to analyse the data on BC collection in the ER before and after a three-phase project. The first phase (1 January to 30 June 2015) before the intervention consisted of evaluation of data on BCs routinely collected in the ER. The second phase (1 July to 31 December 2015) was the intervention phase in which educational courses on sepsis recognition and on pre-analytical phase procedures (including direct incubation) were provided to ER staff. The third phase (1 January to 30 June 2016; after the intervention) again consisted of evaluation. Before the intervention, out of 24,738 admissions to the ER, 103 patients (0.4%) were identified as septic and had BCs drawn (359 BC bottles); 19 out of 103 patients (18.4%) had positive BCs. After the intervention, out of 24,702 admissions, 313 patients (1.3%) had BCs drawn (1,242 bottles); of these, 96 (30.7%) had positive BCs. Comparing the first and third periods, an increase in the percentage of patients with BCs collected (from 0.4% to 1.3% respectively, p < 0.0001) and an increase in the percentages of patients with true-positive BCs (from 0.08% to 0.39% of all patients evaluated respectively, p < 0.0001) were observed. The isolation of bacteria by BCs increased 3.25-fold after project implementation. These results can be principally ascribed to an improved awareness of sepsis in the staff associated with improved pre-analytical phase procedures in BC collection.