Spread of Carbapenem-Resistant Klebsiella pneumoniae in Hub and Spoke Connected Health-Care Networks: A Case Study from Italy.

The study describes the spread of carbapenem-resistant Klebsiella pneumoniae (CRKP) in a regional healthcare network in Italy. The project included several stages: (1) Establishment of a laboratory-based regional surveillance network, including all the acute care hospitals of the Marches Region (n = 20). (2) Adoption of a shared protocol for the surveillance of Multi-Drug Resistant Organisms (MDROs). Only the first CRKP isolate for each patient has been included in the surveillance in each hospital. The anonymous tracking of patients, and their subsequent microbial records within the hospital network, allowed detection of networks of inter-hospital exchange of CRKP and its comparison with transfer of patients within the hospital network. Pulsed-Field Gel Electrophoresis (PFGE) analysis has been used to study selected isolates belonging to different hospitals. 371,037 admitted patients have been included in the surveillance system. CRKP has shown an overall incidence rate of 41.0 per 100,000 days of stay (95% confidence interval, CI 38.5-43.5/100,000 DOS), a CRKP incidence rate of isolation in blood of 2.46/100,000 days of stay (95% CI 1.89-3.17/100,000 days of stay (DOS) has been registered; significant variability has been registered in facilities providing different levels of care. The network of CRKP patients' exchange was correlated to that of the healthcare organization, with some inequalities and the identification of bridges in CRKP transfers. More than 73% of isolates were closely related. Patients' exchange was an important route of spread of antimicrobial resistance, highlighting the pivotal role played by the hub, and selected institution to be used in prioritizing infection control efforts.

Detection of viable but non-culturable Pseudomonas aeruginosa in cystic fibrosis by qPCR: a validation study.

BACKGROUND: Routine culture-based diagnosis of Pseudomonas aeruginosa lung infection in Cystic Fibrosis (CF) patients can be hampered by the phenotypic variability of the microorganism, including its transition to a Viable But Non-Culturable (VBNC) state. The aim of this study was to validate an ecfX-targeting qPCR protocol developed to detect all viable P. aeruginosa bacteria and to identify VBNC forms in CF sputum samples. METHODS: The study involved 115 P. aeruginosa strains of different origins and 10 non-P. aeruginosa strains and 88 CF sputum samples, 41 Culture-Positive (CP) and 47 Culture-Negative (CN). Spiking assays were performed using scalar dilutions of a mixture of live and dead P. aeruginosa ATCC 9027 and a pooled P. aeruginosa-free sputum batch. Total DNA from sputum samples was extracted by a commercial kit, whereas a crude extract was obtained from the broth cultures. Extracellular DNA (eDNA) interference was evaluated by comparing the qPCR counts obtained from DNase-treated and untreated aliquots of the same samples. The statistical significance of the results was assessed by the Wilcoxon test and Student's t test. RESULTS: The newly-developed qPCR protocol identified 96.6% of the P. aeruginosa isolates; no amplification was obtained with strains belonging to different species. Spiking assays supported protocol reliability, since counts always matched the amount of live bacteria, thus excluding the interference of dead cells and eDNA. The protocol sensitivity threshold was 70 cells/ml of the original sample. Moreover, qPCR detected P. aeruginosa in 9/47 CN samples and showed higher bacterial counts compared with the culture method in 10/41 CP samples. CONCLUSIONS: Our findings demonstrate the reliability of the newly-developed qPCR protocol and further highlight the need for harnessing a non-culture approach to achieve an accurate microbiological diagnosis of P. aeruginosa CF lung infection and a greater understanding of its evolution.

Candidemia in the elderly: What does it change?

BACKGROUND: Candidemia is a life-threatening fungal infection and it can affect patients of all ages. Characterization of candidemia in the elderly is lacking. METHODS: We performed a retrospective study of adults (≥ 18 years) with candidemia diagnosed in our center in 2010-2015. Demographics, comorbidities, clinical and microbiologic characteristics, antifungal treatment and outcome were compared between older (≤65 years) and younger (>65 years) patients. RESULTS: Among 302 patients with candidemia identified during the study period, 188 (62%) belonged to the elderly group. Comorbidities were significantly more frequent in older patients and included chronic pulmonary diseases, cardiovascular diseases, diabetes mellitus, and chronic renal failure (p ranging from <0.0001 to 0.017). A significantly higher proportion of older patients had septic shock (p = 0.040) at the time of candidemia. Candida albicans accounted for 53% of isolates and there were no significant differences between patients' age and Candida species. Thirty-day mortality was significantly higher in older (45%) than in younger (28%) patients (p = 0.003). Factors associated with a significant higher proportion of death in the elderly included older age (i.e.: old-old), being hospitalized in ICU rather than in other wards, suffering from chronic pulmonary diseases, the presence of septic shock, multiple organ failure, dialysis and being infected with C. glabrata (p ranging from <0.0001 to 0.034). On multivariate analysis septic shock (HR 1.744 [CI95% 1.049-2.898], p = 0.032) and multiple organ failure (HR 2.242 [CI95% 1.070-4.698], p = 0.032) were independently associated with a higher risk of death. The probability of 30-days survival of older patients was significantly reduced when compared to that of younger patients (p = 0.005) who did not receive any treatment. In the elderly, there was a trend toward higher MICs for fluconazole/C. albicans, fluconazole/C. glabrata, amphotericin B/C. albicans, and caspofungin/C. glabrata. CONCLUSIONS: In our study, we found that elderly patients with Candida bloodstream infections are characterized by a high mortality rate. In particular, the lack of any antifungal therapy as well as the occurrence of septic shock increased significantly the overall mortality. Additionally, we found that there was a trend of higher MIC for specific drug/Candida combination.

Carbapenem-Resistant Klebsiella pneumoniae influences the outcome of early infections in liver transplant recipients.

BACKGROUND: Infections remain a leading cause of morbidity and mortality among liver transplant (LT) recipients. The aim of our study was to define the factors associated with outcome of early bacterial and fungal infections in a cohort of patients who underwent LT at the University Hospital of Ancona over a nine year period. METHODS: All consecutive patients who underwent LT in our center were considered. An early infection was defined as occurring in the first month post-transplantation. RESULTS: Among 330 patients who underwent LT from August 2005 to October 2014, 88 (27 %) had at least one infection documented within 30 days after transplantation. In 54 cases only one site was involved, in 34 cases ≥2 sites. There were 43 (30 %) pneumonia, 40 (27 %) surgical site infections, 31 (22 %) blood stream infections, and 30 (21 %) urinary tract infections. Gram-negative bacteria accounted for 64 % of the culture-positive cases, followed by Gram-positive bacteria (30 %) and fungi (6 %). A high proportion of drug-resistant strains was found within either Gram-negative (79 %) or Gram-positive (81 %) bacteria. There were 27 out 88 patients (31 %) who died within 180 days from the transplant. Factors independently associated with a higher risk of mortality were: renal replacement therapy (HR 11.797 [CI95 % 3.082-45.152], p < 0.0001), multisite infections (HR 4.865 [CI95 % 1.417-16.700], p = 0.012) and being infected with carbapenem-resistant Klebsiella pneumoniae (CRKP; HR 5.562 [CI95 % 1.186-26.088], p = 0.030). CONCLUSIONS: Overall, these data indicate that early infections in LT patients are characterized by significant mortality. In particular, an early infection caused by CRKP has an adverse impact on survival in these patients suggesting an urgent need for adopting preventive measures to avoiding this complication.

Factors related to outcome of bloodstream infections due to Candida parapsilosis complex.

BACKGROUND: Although Candida albicans is the most common cause of fungal blood stream infections (BSIs), infections due to Candida species other than C. albicans are rising. Candida parapsilosis complex has emerged as an important fungal pathogen and became one of the main causes of fungemia in specific geographical areas. We analyzed the factors related to outcome of candidemia due to C. parapsilosis in a single tertiary referral hospital over a five-year period. METHODS: A retrospective observational study of all cases of candidemia was carried out at a 980-bedded University Hospital in Italy. Data regarding demographic characteristics and clinical risk factors were collected from the patient's medical records. Antifungal susceptibility testing was performed and MIC results were interpreted according to CLSI species-specific clinical breakpoints. RESULTS: Of 270 patients diagnosed with Candida BSIs during the study period, 63 (23 %) were infected with isolates of C. parapsilosis complex which represented the second most frequently isolated yeast after C. albicans. The overall incidence rate was 0.4 episodes/1000 hospital admissions. All the strains were in vitro susceptible to all antifungal agents. The overall crude mortality at 30 days was 27 % (17/63), which was significantly lower than that reported for C. albicans BSIs (42 % [61/146], p = 0.042). Being hospitalized in ICU resulted independently associated with a significant higher risk of mortality (HR 4.625 [CI95% 1.015-21.080], p = 0.048). Conversely, early CVC removal was confirmed to be significantly associated with a lower risk of mortality (HR 0.299 [CI95% 0.102-0.874], p = 0.027). Finally, the type of primary antifungal therapy did not influence the outcome of infection. CONCLUSIONS: Candidemia due to C. parapsilosis complex, the second most commonly causative agent of yeast BSIs in our center, is characterized by a non-negligible mortality at 30 days. An early CVC removal is associated with a significant reduced mortality.

Ongoing outbreak of invasive listeriosis due to serotype 1/2a Listeria monocytogenes, Ancona province, Italy, January 2015 to February 2016.

In the first seven weeks of 2016, five serotype 1/2a Listeria monocytogenes isolates were collected from patients with invasive listeriosis in Ancona province in Italy. These strains and six 1/2a isolates identified in 2015 in the same area were typed by ERIC-PCR and PFGE. A clonal relationship, documented between the two sets of isolates, suggested a listeriosis outbreak in Ancona that started most probably in 2015. Investigation into the source of infection is still ongoing.

Delay of antifungal therapy influences the outcome of invasive aspergillosis in experimental models of infection.

OBJECTIVES: The aim of the present study was to evaluate the effects of delayed antifungal therapy on the outcome of invasive aspergillosis due to Aspergillus fumigatus in experimental models of infection. METHODS: A clinical isolate of A. fumigatus susceptible to amphotericin B (MIC 0.5 mg/L) and micafungin [minimum effective concentration (MEC) 0.03 mg/L] was used in all experiments. Two models of infection were investigated in immunosuppressed mice: disseminated infection and pulmonary infection. Twenty-four hours (early therapy) and 48 h (delayed therapy) post-infection, the mice were given vehicle, liposomal amphotericin B, micafungin or liposomal amphotericin B plus micafungin (combination). Drug efficacy was assessed by either survival or tissue burden experiments. RESULTS: In disseminated infection, any drug regimen given early significantly prolonged survival. When therapy was delayed, only micafungin and the combination were effective. In pulmonary infection, although there was a trend towards a prolongation of survival of mice treated early with liposomal amphotericin B, only the combination was effective. Similarly, when therapy was delayed, only the combination was effective. In disseminated infection, any drug regimen given early was effective at reducing the cfu in kidney tissue. In pulmonary infection, only liposomal amphotericin B and the combination given early were effective at reducing the cfu in lung tissue. Conversely, when therapy was delayed, no regimen was effective at reducing the tissue burden, regardless of the type of infection. CONCLUSIONS: Our data indicate that delayed initiation of antifungal therapy is deleterious in experimental models of invasive aspergillosis. A combination regimen seems to have some advantages over a single-drug approach when the therapy is started late.

Risk Factors and Impact on Clinical Outcome of Multidrug-Resistant Acinetobacter Baumannii Acquisition in Cardiac Surgery Patients.

OBJECTIVES: Acinetobacter baumannii recently has emerged as an important nosocomial pathogen. The aim of this study was to assess the impact on mortality of multidrug-resistant A. baumannii (MDR-AB) infection/colonization in patients undergoing cardiac surgery and to investigate microbiologic characteristics, epidemiologic spread of this pathogen, and the relative containment measures. DESIGN: Single-center, retrospective cohort study of prospectively collected data. SETTING: Cardiac surgery tertiary-care center. PARTICIPANTS: Patients with positive MDR-AB cultures from September 1, 2009 to December 31, 2011. INTERVENTIONS: Bivariate and multivariate analyses were performed to individualize the risk factors for MDR-AB-infections in cardiac surgery patients. To evaluate the MDR-AB attributable mortality, a retrospective matched cohort study was performed. Incidence density ratio (IDR) was calculated to compare the MDR-AB infection/colonization before and after the introduction of preventive measures adopted following the first cases. MEASUREMENTS AND MAIN RESULTS: MDR-AB acquisition occurred in 14 patients (0,6%) of 2385 patients. At the multivariate analyses, preoperative use of inotropic drugs (OR 18.2, 95% CI 4.6-71.9) and logistic EuroSCORE (OR 1.09, 95% CI 1.06-1.13) were found as independent risk factors. Patients with MDR-AB had 57% cumulative in-hospital mortality; no statistical differences in mortality were observed in the matched group. IDR revealed a significantly decreased incidence of infection/colonization (0.3 per 1,000 days of stay compared with 0.03/1,000 days of stay, p = 0.0001) after the containment measures became effective. CONCLUSIONS: Sicker patients are more susceptible to be infected by A. baumannii, but mortality is not significantly higher compared with other patients with similar characteristics. Adequate measures are fundamental to control the spread of the infection.

Epidemiology, clinical characteristics, and outcome of candidemia in a tertiary referral center in Italy from 2010 to 2014.

PURPOSE: We evaluated the epidemiology, clinical characteristics and outcome of candidemia in a single institution from 2010 to 2014. METHODS: A retrospective observational study of all cases of candidemia was carried out at a University Hospital in Central Italy including five intensive care units (ICUs), 11 medical and 11 surgical wards. Data regarding demographic characteristics and clinical risk factors were collected from the patient's medical records. Antifungal susceptibility testing was performed and MIC results were interpreted according to species-specific clinical breakpoints. RESULTS: A total of 270 episodes of candidemia were identified. Overall incidence rate was 1.5 episodes/1000 hospital admissions. Although Candida albicans represented the most commonly isolated species, its percentage significantly decreased from 68 to 48 % (p = 0.040). The overall 30-day mortality was 35 %. The variables independently associated with a significant higher risk of mortality were: older age; being hospitalized in ICU or in medical wards vs surgical wards; being infected with C. albicans vs other species; the occurrence of septic shock, pneumonia and acute renal failure; the presence of a solid organ tumor or a chronic pulmonary disease. Conversely, an appropriate treatment was confirmed to be significantly associated with a lower risk of mortality. The overall resistance was low and it was noted only among triazoles. CONCLUSIONS: Our study shows that candidemia is a significant source of morbidity and mortality. The identification of risk factors associated with mortality along with the knowledge of local susceptibility may lead to a better management in terms of preventive and therapeutic measures.

The marine environment as a reservoir of enterococci carrying resistance and virulence genes strongly associated with clinical strains.

To gain insights into the relationships and the genetic exchange among environmental and clinical enterococci, 59 strains (29 from marine aquaculture sites and 30 from clinical settings) resistant to tetracycline, erythromycin, ampicillin and/or gentamicin were analysed for the antibiotic resistance tet(M), tet(L), tet(O), erm(A), erm(B), mef blaZ, aac(6')-Ie aph(2″)-Ia and virulence gelE, cylB, efaA and esp genes, and for the copper resistance gene tcrB. Antibiotic resistance and virulence genes were detected more frequently in clinical than in environmental enterococci; the opposite was true for copper resistance. Conjugation experiments demonstrated the transfer of antibiotic resistance genes from marine to clinical enterococci in interspecific mating and the uncommon joint transfer of tet(L) and erm(B). Enterobacterial repetitive intergenic consensus polymerase chain reaction typing evidenced a cluster (90% similarity) encompassing strains carrying multiple antibiotic resistance genes from both sets; the others marine isolates exhibited polyclonality and bore tcrB. Our results demonstrate that antibiotic-resistant marine enterococci bear antibiotic resistance genes transferable to humans and suggest that copper resistance, not observed among clinical strains, may be useful for survival in the environment, whereas virulence genes likely confer no advantage to enterococcal populations adapted to a lifestyle outside the host.

Effects of amphotericin B on Aspergillus flavus clinical isolates with variable susceptibilities to the polyene in an experimental model of systemic aspergillosis.

OBJECTIVES: The aim of the present study was to evaluate the effects of amphotericin B (AMB) on clinical isolates of Aspergillus flavus. METHODS: MICs of both standard AMB and liposomal AMB (L-AMB) were determined using a broth dilution method for seven isolates of A. flavus. AMB MICs were also determined using the Etest. The activity of the polyene was then investigated in a murine model of systemic aspergillosis in which animals were infected intravenously, treated intravenously with several doses of the polyene (1-10 mg/kg/day) and observed for survival. RESULTS: Broth dilution AMB, broth dilution L-AMB and Etest AMB MICs ranged from 0.5 to 2.0 mg/L, 0.06 to >16 mg/L and 1.0 to >32 mg/L, respectively. There were two isolates for which all doses were effective at prolonging the survival. Their AMB MICs were ≤1.0 mg/L, regardless of the method/drug formulation utilized for testing. There were four isolates for which no regimen was effective. Their broth dilution AMB, broth dilution L-AMB and Etest AMB MICs ranged from 1.0 to 2.0 mg/L, 0.06 to >16 mg/L and 2.0 to >32 mg/L, respectively. There was one isolate for which only L-AMB given at 10 mg/kg/day was effective; broth dilution MICs of AMB and L-AMB were 0.5 mg/L, while the Etest MIC of AMB was 2.0 mg/L. CONCLUSIONS: Our data indicate that not all isolates of A. flavus should be considered resistant to AMB. The Etest represented the in vitro method that best correlated with the experimental infection. Finally, a clinical isolate showing an MIC ≥2.0 mg/L may be reasonably considered resistant in vivo to any dose/formulation of the polyene.

In vitro and in vivo effects of echinocandins against Candida parapsilosis sensu stricto, Candida orthopsilosis and Candida metapsilosis.

OBJECTIVES: The aim of the present study was to compare, in vitro and in vivo, the effects of caspofungin, micafungin and anidulafungin against Candida parapsilosis complex isolates. METHODS: In vitro activities of all three echinocandins were assessed against C. parapsilosis sensu stricto (n = 4), Candida orthopsilosis (n = 4) and Candida metapsilosis (n = 3) using broth microdilution susceptibility testing, minimum fungicidal concentration determination and a killing-curve assay, in the absence and in the presence of 50% human serum. Then, the activities of all drugs were investigated in an immunocompromised murine model of systemic candidiasis. Animals were infected with six isolates (two for each species) and treated with the echinocandins administered at 0.25, 1, 5 and 10 mg/kg/day for six consecutive days. Fungal burdens were assessed in kidney tissues on day 7 post-infection. RESULTS: Geometric mean MICs of caspofungin, micafungin and anidulafungin for C. parapsilosis sensu lato were, respectively, 0.09, 0.14 and 0.20 mg/L without serum, and 0.70, 3.92 and 5.84 mg/L with serum. The fungicidal activity of all three echinocandins was variable; however, the addition of serum reduced the fungicidal effects against these species. In vivo studies showed that caspofungin at 5 and 10 mg/kg/day significantly decreased the kidney burdens with respect to the controls for all isolates, while micafungin was active at 5 and/or 10 mg/kg/day only against C. metapsilosis. CONCLUSIONS: Our susceptibility testing showed that caspofungin was the most active echinocandin against all three species. Also, caspofungin resulted in significant therapeutic effects for treatments of experimental systemic infections due to the three species, while micafungin was effective only against C. metapsilosis.

Evaluation of the peptide nucleic acid fluorescence in situ hybridisation technology for yeast identification directly from positive blood cultures: an Italian experience.

Fungaemia is an increasing nosocomial pathology. The 'gold standard' for detection of fungaemia is blood culture, but it is time-consuming and its sensitivity for early detection is low. On the other hand, yeasts present different antifungal sensitivity patterns to be quickly detected to allow an effective treatment. The aim of this study was to evaluate the diagnostic performances of PNA-FISH to directly identify yeasts from blood cultures and to compare results with those obtained by culture. A total of 176 blood cultures positive for yeasts at direct Gram stain and 24 negative blood cultures as control collected from 15 Italian hospitals, included in a network coordinated by the Medical Mycology Committee, Italian Society of Clinical Microbiology (AMCLI), were examined both by culture and PNA-FISH technology. Sensitivity of the PNA-FISH technique evaluated for five Candida species was 99.3% and specificity, 100%. Distinguishing which yeast is implicated in fungaemia and whether the infection is caused by multiple species are important for the selection of antifungal therapy. The PNA-FISH technique is a very useful approach because the test discriminates between groups of Candida species with different susceptibility pattern, particularly against azoles and echinocandins, with only a 90-minute turn-around time after the Gram-stain reading.

Microarray technology for yeast identification directly from positive blood cultures. A multicenter Italian experience.

The authors evaluated the performance of the MycArray™ Yeast ID (Myconostica Ltd, UK) assay in the identification of a total of 88 yeast isolates recovered in culture as compared to that obtained through routine methods. The turn-around time for species identification directly from cultures by the MycArray was 6 hours, much quicker than classical methods and all yeasts were correctly identified. In two cases a double identification including Saccharomyces cerevisiae was noted, but it was not confirmed by culture. The results show that MycArray Yeast ID can be a potential tool for rapid detection and identification of Candida species.

Comparative effects of micafungin, caspofungin, and anidulafungin against a difficult-to-treat fungal opportunistic pathogen, Candida glabrata.

The aim of this study was to compare the in vitro and in vivo activities of micafungin, caspofungin, and anidulafungin against Candida glabrata. The MICs against 28 clinical isolates showed that the overall susceptibilities to caspofungin and to micafungin were not statistically different in the absence of human serum, whereas the isolates were less susceptible to micafungin than to caspofungin in its presence. Minimum fungicidal concentrations, as well as time-kill experiments, showed that caspofungin was more active than anidulafungin, while micafungin was superior to either caspofungin or anidulafungin without serum; its addition rendered caspofungin and micafungin equally effective. A murine model of systemic candidiasis against a C. glabrata-susceptible isolate was performed to study the effects of all three echinocandins, and kidney burden counts showed that caspofungin, micafungin, and anidulafungin were active starting from 0.25, 1, and 5 mg/kg of body weight/day, respectively. Two echinocandin-resistant strains of C. glabrata were selected: C. glabrata 30, a laboratory strain harboring the mutation Fks2p-P667T, and C. glabrata 51, a clinical isolate harboring the mutation Fks2p-D666G. Micafungin activity was shown to be as effective as or more effective than that of caspofungin or anidulafungin in terms of MICs. In vivo studies against these resistant strains showed that micafungin was active starting from 1 mg/kg/day, while caspofungin was effective only when administrated at higher doses of 5 or 10 mg/kg/day. Although a trend toward colony reduction was observed with the highest doses of anidulafungin, a significant statistical difference was never reached.

Interlaboratory evaluation of VITEK2 system and Sensititre YeastOne® for antifungal susceptibility testing of yeasts isolated from blood cultures against four antifungal agents.

An interlaboratory evaluation (seven centers) of VITEK2 System and Sensititre YeastOne® was conducted to test the antifungal susceptibilities of yeasts. The MICs of amphotericin B, fluconazole, flucytosine, and voriconazole were determined for 70 isolates of Candida spp. Our results demonstrated a higher interlaboratory agreement of VITEK 2 System than Sensititre YeastOne©. A good concordance between the two methods was observed for amphotericin B, fluconazole, voriconazole and 5-fluorocytosine (from 81.4% to 88.6%). The study suggests the potential value of the VITEK2 System as a convenient alternative method for testing the susceptibility of yeasts. It also indicates the need for further optimization of MIC endpoint criteria to improve interlaboratory agreement.

Heterogeneity of Tn5253-like composite elements in clinical Streptococcus pneumoniae isolates.

Several drug resistances in Streptococcus pneumoniae are associated with mobile genetic elements, which are loosely subdivided into a group of smaller (18- to 27-kb) and a group of larger (>50-kb) elements. While the elements of the former group, which typically carry the tetracycline resistance determinant tet(M) and whose prototype is Tn916 (18 kb), have been studied extensively, the larger elements, whose prototype is Tn5253 (∼65.5 kb), are not as well explored. Tn5253 is a composite structure consisting of two independent conjugative transposons, Tn5251 (which is virtually identical to Tn916) and Tn5252 (∼47.5 kb), with the former inserted into the latter. Tn5252, which so far has only partially been sequenced, carries an integrase gene, driving its site-specific insertion into the host cell genome, and the chloramphenicol resistance cat(pC194) determinant. This study investigated 20 clinical isolates of S. pneumoniae, which were selected on the basis of cat(pC194)-mediated chloramphenicol resistance. All 20 isolates harbored a Tn5253-like element. The composite elements (some of which have been completely sequenced) demonstrated considerable heterogeneity that stemmed from a dual variability: in the Tn5252-like element, due primarily to differences in the integrase gene but also to differences in cargo genes and in the overall genetic organization, and in the Tn916-like element, with the possible involvement, besides Tn916, of a number of Tn916 family pneumococcal elements carrying different erythromycin resistance genes. In mating experiments, only one composite element, containing a less typical Tn916 family element, appeared to be nonmobile. Being part of a Tn5253-like composite element may confer on some Tn916-like transposons, which are apparently nontransferable as independent genetic elements, the ability to be mobilized.