Ceftazidime/avibactam-resistant meropenem-susceptible KPC-producing Klebsiella pneumoniae: Analysis of cases and evaluation of in vitro activity of fosfomycin-containing combinations.

OBJECTIVES: Little is known regarding outcomes and optimal therapeutic regimens of infections caused by Klebsiella pneumoniae carbapenemase-producing Klebsiella pneumoniae (KPC-Kp) resistant to ceftazidime/avibactam (CZA) and susceptible to meropenem (MEM). Although susceptible to MEM in vitro, the possibility of developing MEM resistance overtime is a concern. We describe the clinical characteristics of patients with colonization/infection due to KPC variants with a focus on the in vitro activity of fosfomycin (FOS)-containing combinations. METHODS: Patients with colonization/infection due to a KPC variant were included. Fosfomycin susceptibility was performed by agar dilution method. Synergistic activity of FOS-based combinations was evaluated by gradient strip-agar diffusion method. The emergence of in vitro MEM resistance was also tested. RESULTS: Eleven patients were included: eight with infection [four with ventilator-associated pneumonia and four with bloodstream infections] and three with colonization. Previous therapy with CZA was administered to all patients (with a mean cumulative duration of 23 days). All subjects with infection received meropenem, in monotherapy (n = 4) or with amikacin (n = 2) or fosfomycin (n = 2), and achieved clinical cure. A new CZA-susceptible and MEM-resistant KPC-Kp strain was subsequently isolated in three patients (27.3%). Meropenem/vaborbactam (MVB) showed high in vitro activity, while FOS+MEM combination was synergistic in 40% of cases. In vitro resistance to MEM was observed with maintenance of CZA resistance. CONCLUSIONS: Detection of KPC variants may occur within the same patient, especially if CZA has been previously administered. Although clinical success has been obtained with carbapenems, the emergence of MEM resistance is a concern. Fosfomycin plus meropenem is synergistic and may be a valuable combination option for KPC variants, while MVB may be considered in monotherapy. The detection of KPC variants in an endemic setting for KPC-Kp represents a worryingly emerging condition. The optimal therapeutic approach is still unknown and the development of meropenem resistance is of concern, which may lead to therapeutic failure in clinical practice. In these cases, the addition of fosfomycin to meropenem, or a more potent antibiotic, such as meropenem/vaborbactam, may be valuable therapeutic options.

Role of Double-Carbapenem Regimen in the Treatment of Infections due to Carbapenemase Producing Carbapenem-Resistant Enterobacteriaceae: A Single-Center, Observational Study.

PURPOSE: (i) To compare infections caused by carbapenem-susceptible (CS) and carbapenemase producing carbapenem-resistant Enterobacteriaceae (CP-CRE); (ii) to evaluate the clinical effectiveness of the double-carbapenem (DC) regimen in comparison with the best available treatment (BAT) in infections caused by CP-CRE; and (iii) to determine the exact minimal inhibitory concentrations (MICs) of meropenem/ertapenem (MEM/ETP) and the degree of in vitro ETP+MEM synergism in subjects receiving the DC. METHODOLOGY: Over a 3-year period (2014-2017), patients with infections due to Enterobacteriaceae were included in a single-center, retrospective, observational study. According to the susceptibility to carbapenems, subjects were divided into CSE and CP-CRE groups. CP-CRE group was further divided into subjects receiving the DC regimen and those treated with other regimens (BAT group). Clinical characteristics and the presence of 5th-day response and 60-day outcome were evaluated for DC and BAT groups. The determination of MEM and ETP actual MICs and the MEM+ETP synergistic activity were performed on strains obtained from subjects receiving the DC regimen. RESULTS: A total of 128 patients were included in the study: 55/128 (43%) with infections due to CP-CRE and 73/128 (57%) with infections due to CSE. Among CP-CRE (n=55), 21 subjects (39%) were treated with the DC regimen whereas 34 (61%) received BAT. No differences in terms of severity of infection, presence/absence of concomitant bacteremia, type of infection, and resolution of infection were found; in contrast, DC group tended to have a higher rate of sepsis or septic shock at the onset of infection and a higher rate of 5th-day response. MICs 50/90 were 256/512 and 256/256 µg/mL for MEM and ETP, respectively. Overall, complete in vitro synergism was found in 6/20 strains (30%). CONCLUSION: The DC regimen is a valid and effective therapeutic option in patients with infections due to KPC producing CRE, including those with bacteremic infection and more severe clinical conditions. The clinical effectiveness is maintained even in the presence of extremely high MEM MICs.

In vitro evaluation of different antimicrobial combinations against carbapenemase-producing Klebsiella pneumoniae: the activity of the double-carbapenem regimen is related to meropenem MIC value.

Objectives and methods: We evaluated the in vitro activity of different antimicrobial combinations with and without colistin against 39 carbapenem-resistant Klebsiella pneumoniae (CR-Kp) strains (colistin + meropenem/doripenem, colistin + tigecycline, colistin + rifampicin, gentamicin + meropenem, gentamicin + tigecycline and the double-carbapenem regimen meropenem + ertapenem) using the chequerboard method. The triple combination colistin + meropenem + tigecycline was also tested. In addition, killing studies were performed for meropenem + ertapenem. Results: Gentamicin-based combinations showed a high level of synergy. Meropenem + ertapenem was synergic in 12/39 (30.7%) of the strains, whereas based on killing studies 1 × MIC meropenem + 1 × MIC ertapenem and 2 × MIC meropenem + 1 × MIC ertapenem combinations were bactericidal and synergic at 24 h [mean area under the bactericidal curve (AUBC) 54.9 ±âŸ26.1 and 44.2 ±âŸ15.3 compared with 1 × MIC meropenem (134.5 ±âŸ40.1) and 2 × MIC meropenem (126.4 ±âŸ5.4), respectively, P < 0.0001]. When the results were stratified according to meropenem MIC, we found that the degree of synergy significantly increased for isolates with lower meropenem (and not ertapenem) MICs, up to an MIC of 128 mg/L. Among colistin-containing combinations, synergy was observed in 18/39 (46.1%), 33/34 (97%), 24/39 (61.5%) and 17/39 (43.5%) of the strains for colistin + meropenem, colistin + rifampicin, colistin + tigecycline and colistin + doripenem, respectively, including colistin-resistant strains. Colistin + meropenem + tigecycline at subinhibitory concentrations resulted in the absence of growth of 37/39 strains (94.8%). Conclusions: Our in vitro data suggest that colistin might be a valid therapeutic option against CR-Kp, even in the presence of colistin resistance, whereas the double-carbapenem regimen represents a viable option when colistin is not recommended, especially if the meropenem MIC is ≤ 128 mg/L. Since traditional antimicrobial susceptibility reports are not sufficiently informative for clinicians, synergy testing as well as actual meropenem MIC evaluation should always be performed in the case of CR-Kp infections.

Bactericidal and synergistic activity of double-carbapenem regimen for infections caused by carbapenemase-producing Klebsiella pneumoniae.

Available therapeutic options against carbapenem-resistant Klebsiella pneumoniae (CR-Kp) are limited because of the high level of resistance to other antimicrobial classes including polymyxins. The double-carbapenem regimen has been recently considered a possible therapeutic strategy. In the present study, we evaluated the in vitro bactericidal and synergistic activity of a double-carbapenem regimen consisting of ertapenem plus high-dose meropenem in a series of patients with healthcare-associated CR-Kp infections in whom the use of colistin was not indicated because of potential nephrotoxicity and/or resistance. In vitro synergy was evaluated using checkerboard and killing studies. A total of 15 patients were included in the study, with sepsis, severe sepsis and septic shock found in two (13.3%), five (33.3%) and one (6.7%) patients, respectively. Overall, the clinical/microbiological response was 12/15 (80%). Synergy was observed in 11/14 (78.6%) isolates using the checkerboard method whereas in killing studies 12/14 (85.7%) and 14/14 (100%) strains were synergistic and bactericidal at 24 h at concentrations of 1 × MIC MEM+1 × MIC ERT and 2 × MEM+1 × MIC ERT, respectively, with a significant decrease of log CFU/mL compared with other combinations (p <0.0001). The double-carbapenem regimen showed clinical and in vitro effectiveness in patients with CR-Kp infections.

Study of methicillin-resistant Staphylococcus aureus (MRSA) carriage in a population of HIV-negative migrants and HIV-infected patients attending an outpatient clinic in Rome.

BACKGROUND: Migration and HIV infection are known risk factors for methicillin-resistant Staphylococcus aureus (MRSA) carriage and infection. The aim of the study was to analyze the prevalence of MRSA nasal colonization in a high risk population of HIV-negative migrants and HIV-infected subjects. Secondary aim was to investigate over time MRSA carriage prevalence in HIV-infected subjects. METHODS: During the study period (January-June 2008), nasal swabs were collected from 96 HIV-negative migrants and 63 HIV-infected patients. A group of 68 seropositive subjects was additionally screened for MRSA carriage in 2012. Subjects were evaluated for HIV status, previous antibiotic use or hospitalization, soft tissue and skin infections (SSI), nationality and work conditions. The swab specimens were plated and incubated for 24-h under static condition at 37 degrees and then identified as S. aureus by using standard methods. RESULTS: A total of 227 subjects, 131 HIV-infected adults (63 in 2008 and 68 in 2012) and 96 HIV-negative migrants, were analyzed. Overall, 71/227 (31.2%) were S. aureus carriers: 34 out of 131 (25.9%) among HIV infected subjects and 37 out of 96 (38.5%) among migrants. Two MRSA were detected in HIV-infected patients (2.8%). Between 2008 and 2012 there was an increase of MRSA carriage in HIV+ group (p=0.49). No statistically significant differences were found between S. aureus carriers and no-carriers in terms of CD4+ cell count, TMP/SMX prophylaxis, previous antibiotic use or hospitalization, nationality and duration of stay in Italy. Among HIV+ patients there was a higher prevalence of SSI in MSSA carriers compared with no carriers (25% vs 4%, p=0.028). In the migrants group, having a job based on a close human contact was significantly associated with S. aureus colonization (p=0.0038). CONCLUSIONS: Despite of the high prevalence of S. aureus isolation (31.2%), the present study showed the low rate of MRSA carriage in a high risk population. The main factor associated with S. aureus colonization was a close human contact rather than the HIV status and the condition of being migrant.

Helicobacter pylori: determinant and markers of virulence.

In this review, we examine the main virulence determinants in Helicobacter pylori (Hp) strains and the correlation between these and the different diseases following Hp infections. We also discuss the host response to Hp and the implications and advantages of the development of non-invasive pre-endoscopy screening of molecular and serological markers associated with Hp virulence and disease progression putting an emphasis on new screening techniques such as the Luminex--X multi analytes profiling technology and the multiplex PCR.

Activity of D-carnitine and its derivatives on Trypanosoma infections in rats and mice.

Little progress has been made in the treatment of African trypanosomiasis over the past decades. L-carnitine has a major role in glycolysis-based energy supply of blood trypanosomes for it stimulates constant ATP production. To investigate whether administration of the isomer D-carnitine could exert a competitive inhibition on the metabolic pathway of the L-form, possibly resulting in parasite replication inhibition, several formulations of this compound were tested on Trypanosoma lewisi and T. brucei rhodesiense in rodent models. High oral dosages of D-cornitine inner salt and proprionyl-D-carnitine were not toxic to animals and induced about 50% parasite growth inhibition in reversible, i.e. competitive, fashion. A putative mechanism could be an interference in pyruvate kinase activity and hence ATP production. Considering both, lack of toxicity and inhibitory activity, D-carnitine may have a role in the treatment of African trypanosomiasis, in association with available trypanocidal drugs.

Reduced bactericidal activity against Staphylococcus aureus and Pseudomonas aeruginosa of blood neutrophils from patients with early adult respiratory distress syndrome.

This study investigated the bactericidal capability of circulating neutrophils from blunt trauma patients admitted to an Intensive Care Unit against Staphylococcus aureus and Pseudomonas aeruginosa. Among those patients, two groups were considered and compared: patients who developed adult respiratory distress syndrome (ARDS) and patients who developed only pneumonia. Peripheral blood samples were drawn as soon as a diagnosis of pneumonia or ARDS was made, followed by the isolation of neutrophil cells and assessment of bacteria phagocytosis and killing. The results demonstrated that in patients with ARDS, phagocytosis and killing efficiency were significantly impaired in comparison with patients with pneumonia and healthy controls. A possible dysregulation of reactive oxygen species production involving the release of humoral mediators in early ARDS may be involved.

Antimicrobial investigation of semipurified fractions of Ginkgo biloba leaves.

A total methanolic extract of Ginkgo biloba leaves was fractionated by solvent partition using ethyl acetate (fraction A), n-butanol (fraction B) and water (fraction C). The antimicrobial activity of the three fractions was evaluated using a number of Gram-positive and -negative bacteria and yeasts. The apolar fraction A appeared to be the most interesting because of its activity against several microorganisms; this fraction was further separated by high performance liquid chromatography, and shown to contain substances with strong inhibitory activity against Enterococcus faecalis 31, different from the major known chemical components of G. biloba leaves.

Antimicrobial activity of fluoroquinolones and other antibiotics on 1,116 clinical gram-positive and gram-negative isolates.

A total of 1,116 clinically isolated strains belonging to Staphylococcus aureus (200), Staphylococcus epidermidis (200), Streptococcus pneumoniae (20), Escherchia coli (200), Klebsiella spp. (177), Serratia marcescens (22), Pseudomonas aeruginosa (224), Haemophilus influenzae (35) and Salmonella (38) from the Department of Infectious Diseases, La Sapienza University in Rome (Italy) were tested against three fluoroquinolones (ofloxacin, ciprofloxacin and levofloxacin) and 10 other antibiotics (augmentin, ampicillin, cefaclor, cefixime, cefotaxime, cotrimoxazole, gentamicin, minocycline, oxacillin and vancomycin). Fluoroquinolones inhibited essentially about 100% of H. influenzae, Salmonella and S. pneumoniae, more than 75% of Staphylococcus including methicillin-resistant strains, and about 90% of Enterobacteriaceae and 50% of P. aeruginosa. Minimal inhibitory concentration values ranged from < 0.015 to > 32 micrograms/ml for Klebsiella, S. aureus and epidermidis, E. coli and P. aeruginosa; from < 0.015 to 2 micrograms/ml for Salmonella; from 0.03 to 16 micrograms/ml for Serratia; from < 0.015 to 1 microgram/ml for Haemophilus; and from 0.5 to 2 micrograms/ml for S. pneumoniae. Levofloxacin and to a lesser extent ofloxacin and ciprofloxacin, generally exhibited a greater activity than the other agents against both Gram-positive and Gram-negative bacteria. Regarding the distribution of resistant strains in Italy, we found a peculiar pattern of resistance as far as E. coli and P. aeruginosa were concerned. Quality control parameters are also summarized. S. epidermidis resulted as a new emergent pathogen especially in immunocompromised patients and its level of sensitivity has been modified over the last few years. In fact, the percentage of resistant strains to antibiotics or the percentage of methicillin-resistant isolates (in our study 35%), has gradually increased. Levofloxacin and ofloxacin showed good activity against staphylococcal strains compared with the majority of other antibiotics. These results suggest that the newer quinolones are promising antimicrobial agents for various infections.

The influence of the SOS response on the activity of 4-quinolones and zidovudine against some strains of Enterobacteria.

The 4-Quinolones are known to induce the SOS response. This should also be the case with AZT (Zidovudine) which has the same bactericidal mechanism. SOS response might make the bacteria more sensitive or more resistant to subsequent doses of quinolones and AZT. NA (Nalidixic acid), the first quinolone of the early 1960s, sensitises a strain of E. coli isolated from the urine of patients with cystopyelitis and the E. coli AB1157 wild type strain which is a well-known SOS inducer. In this case, the SOS system is not involved but only the recombination repair mechanisms which make the bacteria more susceptible to further damage by NA. On the contrary, CPX (Ciprofloxacin) protects E. coli from further exposure to antibiotics. Therefore the SOS response induction assists the bacteria in recovering from the DNA damage caused by CPX. The SOS response induced by AZT in the tested E. coli strains does not seem to either contribute to the lethality of the drug or to be involved in protecting bacteria from the damage caused by AZT. In fact, the percentage of killing was the same for both pre-treated and non pre-treated bacteria (p = 0.5). On the contrary, it was found that in Salmonella typhimurium belonging to blood of a patient with recurrent bacteriaemia, the CPX added to pre-treated bacteria with AZT was less lethal than when it was added to non pre-treated bacteria. The SOS response, in this case, protects bacteria from the damage caused by AZT.

["Killing" of pseudomonas aeruginosa isolated from patients with critical post-operative complications. Effect of various antibiotics]. / "Killing" di pseudomonas aeruginosa isolati da pazienti con complicazioni critiche post-operatorie. Influenza di alcuni antibiotici.

We undertook this study to estimate phagocytic killing by neutrophils (PMNs) of Pseudomonas aeruginosa pre-exposed to sub-inhibitory concentration of Amikacin and Imipenem. In particular, we have isolated bacteria from endotracheal aspirates of post-operative patients mechanically ventilated admitted to an ICU with respiratory failure. PMNs were obtained both from these patients (Group A, n. 6) as well as from subjects submitted to surgery with uncomplicated post-operative period (Group B, n. 8). From specimens tested, 6 strains of Pseudomonas aeruginosa were isolated. Results showed that the rate of killing of bacteria treated with Amikacin was no different from that of untreated bacteria, whichever the source of PMNs, either from Group A or Group B patients. On the other hand, the microbicidal effect on P. aeruginosa exposed to Imipenem was significantly enhanced when PMNs were obtained from Group B patients. In the mixture bacteria, Imipenem and PMNs obtained from Group A the rate of killing was low, similar to the controls without antibiotics. Such a finding suggests a possible impairment of PMNs due to the critical disease and in some way responsible for the host adverse interaction between granulocytes, antibiotics and pathogens. The underlying mechanisms remain to be clarified and further studies are required to understand the possible clinical implications.

Incidence of bacterial colonization in the throat and in urines at paediatric age with evaluation of sensitivity to common antibiotics.

OBJECTIVES: To evaluate the incidence of bacterial colonization in the throat and in urines of children admitted to a paediatric ward in the year 1994. To test the sensitivity of isolates on the most common antibiotics used in therapy. METHODS: The investigation was carried out on a group of 270 children (125 male and 145 female), aged between 3 months and 12 years, hospitalized with feverish infectious pathology in the department of infectious and Tropical Diseases of the University "La Sapienza" of Rome. The cultures of the throat swabs and on urines were performed on the admission of the children before the beginning of the therapy. RESULTS: The throat-swab cultures showed pathogenous microrganisms in 232 samples (85.9%) with a slight prevalence of Gram-negative bacteria (122) with respect to Gram-positive (110) and saprophytic microbial flora (38). The urine cultures proved to be positive in 81 cases (30%) with a prevalence of Gram-negative (56) above Gram-positive isolates (25). CONCLUSIONS: The two/thirds of paediatric patients hospitalized in an Infectious Diseases Department appeared to be colonized in the upper respiratory tract, whereas in about 10% of them a marked bacteriuria was clearly evident, often in the absence of specific symptoms. A few isolates either from the throat or from urines, showed resistance to the common antibacterial agents.

Type frequency and antimicrobial susceptibility of Mycobacterium avium-intracellulare complex strains isolated in Italy from AIDS and non-AIDS patients.

Typing of the glycopeptidolipid antigens performed by thin layer chromatography on 59 Mycobacterium avium-intracellulare (MAC) strains isolated in Italy from AIDS patients showed that the most frequent types were 1, 4, 3, 8, and 21 (24, 19, 14, 14 and 8% of the strains, respectively). Among non-AIDS patients, types 1, 4 and 8 were also frequently found. The antimicrobial susceptibility tested in agar and/or liquid media to a panel of drugs indicated in clofazimine and rifabutin effective agents against both AIDS and non-AIDS strains. The data obtained show that MAC type distribution in Italy appears to be different from that reported for other countries. No major differences in drug susceptibility between AIDS and non-AIDS related strains were found.

Activity of seven antimicrobial agents, alone and in combination, against AIDS-associated isolates of Mycobacterium avium complex.

The activity of seven antimicrobial agents (and five two-drug combinations and five three-drug combinations) was investigated against 37 clinical isolates of Mycobacterium avium recovered from blood cultures of AIDS patients. The susceptibility tests were performed in Middlebrook 7H12 broth using a radiometric method. MICs of amikacin, ciprofloxacin, clarithromycin, clofazimine, ethambutol, rifabutin and sparfloxacin were determined. Five antimicrobial agents were tested in combination with clarithromycin and also with clarithromycin plus amikacin to look for possible synergic activity. Synergic activity in combination with clarithromycin and with clarithromycin plus amikacin, was detected for rifabutin (54% and 51% of isolates, respectively), clofazimine (38% and 35%), ethambutol (16% and 32%), ciprofloxacin (8% and 14%) and sparfloxacin (3% and 8%). No antagonism was observed. We conclude that clarithromycin is an essential component in the chemotherapy of M. avium complex disease.