The aim of this study is to analyze ozone impact on some cariogenic bacteria in ex vivo and in vitro conditions. The in vitro part of study inoculated dentine with strains of Streptococcus mutans ATCC 33402 and Lactobacillus paracasei ATCC 11974 bacteria. Samples of dentine before and after 40s ozone treatment were collected and anaerobically incubated. Samples of cariogenic dentine (N = 24) were collected from permanent molars within the ex vivo segment of the study, prior and after 40s ozone treatment and a number of colonies were counted after incubation. For the in vitro part of study, results have shown a statistically significant average value of reduction of Streptococcus mutans ATCC 33402 and Lactobacillus paracasei ATCC 11974 prior and after ozone treatment (p < 0.001). The ex vivo segment of the study has also demonstrated a statistically significant difference in the number of bacteria prior and after ozone implementation (p < 0.001). Gaseous ozone demonstrated a strong antimicrobial effect on cariogenic bacteria in both in vitro and ex vivo conditions and it can be used as an adjuvant in caries therapy.

The aim of this retrospective study was to evaluate and compare the incidence, timing and etiology of bloodstream infections (BSIs) in patients treated with liver-(LT) or hematopoietic stem cell transplantation (HSCT) in a single institution. We evaluated 280 consecutive transplantations over a period of 34 months. Our results demonstrated 84 episodes of BSIs (47 in LT patients and 37 in HSCT patients) at a median of 28 days post-transplantation. Relative incidence of 34.6 and 29.4 BSI episodes per 100 LT and HSCT patients, respectively, did not differ significantly between the two groups (p = 0.52). BSIs in HSCT patients occurred significantly earlier (p = 0.003) than in LT patients. The recently described reemergence of gram-negative (GN) pathogens as causative agents of BSIs in these patients was confirmed: GN bacilli were the predominant isolates in the LT group, responsible for 58.5% of BSIs and a very frequent (39%) cause of BSIs in the HSCT group. A higher incidence of resistant enterobacteriaceae producing extended spectrum beta-lactamases was found in isolates from LT patients compared to HSCT patients. In both groups, Pseudomonas aeruginosa was the most difficult to treat organism, with 57% of these isolates in LT patients and 44% in HSCT patients being resistant to carbapenems. To conclude, BSIs were confirmed to be important infectious complications of both LT and HSCT. Surveillance and analysis of bacteria causing bloodstream and other serious infections in transplanted patients remain the main prerequisites for planning interventions regarding prevention and treatment of infections in these patients.

We read with great interest the article by Bert et al.1 about the microbial epidemiology of bloodstream infections (BSIs) after liver transplantation (LT). We would like to comment on this issue on the basis of our results, which support the reemergence of gram-negative (GN) bacteria as a leading cause of BSIs after LT. The use of antibacterial and antifungal prophylaxis through the decades has likely contributed to the emergence of increasingly resistant pathogens that now necessitate alterations in antimicrobial therapy. After the introduction of selective digestive decontamination against GN microorganisms in the mid 1990s, gram-positive (GP) bacteria emerged as major pathogens. However, Singh et al.2 in 2004 and now Bert et al.1 have reported the reemergence of GN bacteremias. We analyzed the occurrence of BSIs in 136 consecutive LT patients (mean age = 49 years) during the first year after LT. The study period was January 2005 to October 2008. We considered BSIs to be significant according to the criteria of the Centers for Disease Control and Prevention: (1) the isolation of a microorganism other than a common skin contaminant in at least 1 blood culture with signs and symptoms of infection or (2) the isolation of a common skin contaminant in at least 2 blood cultures drawn on separate occasions with signs and symptoms of infection.3 Blood cultures were incubated with the BacT/ALERT automated system (BioMerieux, France). Isolates were identified with standard microbiology procedures, and sensitivity testing was performed by disk diffusion according to the standards and interpretation of the Clinical and Laboratory Standards Institute.4 Minimal inhibitory concentrations were determined with Etest (AB Biodisk, Sweden). All patients were treated with perioperative prophylaxis (piperacillin-tazobactam for 5 days) and selective digestive decontamination (gentamicin and fluconazole for 3 days). Standard prophylaxis with trimethoprim-sulfamethoxazole (9 months) for Pneumocystis jiroveci and with valganciclovir (3 months) for cytomegalovirus was given to all patients. Immunosuppression consisted of prednisone induction and a maintenance regimen: prednisone (withdrawal after 3-6 months), calcineurin inhibitors (tacrolimus or cyclosporine), and mycophenolate mofetil. Forty-seven BSIs were identified in 23 transplant patients (16.9%); this represents an incidence of 34.6 episodes per 100 transplant patients. They occurred predominantly in the first trimester (median = 46 days post-LT, range = 0-248 days, standard deviation = 72 days). Polymicrobial bacteremias accounted for 12.8% of cases. Altogether, 53 pathogens were isolated. GN bacilli (predominantly Klebsiella pneumoniae, Pseudomonas aeruginosa, and Escherichia coli) were responsible for 58.5% of BSIs (Table 1). A high incidence (42%) of resistant Enterobacteriaceae producing extended-spectrum beta-lactamases (ESBLs) was observed among BSI isolates (Table 2). Low sensitivity in Enterobacteriaceae was recorded for piperacillin/tazobactam (63%), third-generation cephalosporins (58%), cefepime (58%), gentamicin (47%), amikacin (53%), and ciprofloxacin (21%). We found a high proportion of P. aeruginosa isolates to be resistant to multiple antibiotics, including carbapenems (57%), in our series of patients. The most prevalent GP microorganisms that were isolated were enterococci and coagulase-negative staphylococci (CoNS). In the CoNS cases, all BSIs were catheter-related according to clinical criteria. Methicillin-resistant Staphylococcus aureus (MRSA) was responsible for only 1 BSI. Fungi accounted for a higher proportion of isolates (13.2%) in comparison with other studies in which antifungal prophylaxis was used.5 Most fungemias were caused by Candida albicans(50%), whereas non-albicans species included Candida parapsilosis and Candida tropicalis, which were fluconazole-susceptible. Death occurred within 15 days as a result of BSIs in 8 of the 47 episodes (17%), and it was predominantly associated with ESBL-producing K. pneumoniae(3 cases), carbapenem-insusceptible P. aeruginosa(3 cases), and Candida parapsilosis(2 cases). The only vancomycin-resistant isolate of Enterococcus faecium(vancomycin-resistant enterococci) resulted in a fatal outcome. In 3 patients who died of BSIs, multiple organisms were isolated from blood cultures. In conclusion, our data support the recently described reemergence of GN pathogens as leading agents of BSIs after LT. It is particularly worrisome that a large proportion of GN bacteria causing BSIs are resistant to multiple antibiotics; this not only compromises the empirical therapy of sepsis in LT patients but also leaves few choices in some cases even when the causative agent is known. Carbapenem-resistant nonfermentative bacteria are becoming more prevalent in our unit, and it may be only a matter of time before colistin is no longer effective as a drug of last resort. Rigorous infection control practices should be applied in an attempt to stop the spread of such organisms once they appear on the ward. No less important is the need to rationalize the use of antibiotics, which in many cases are misused in the treatment of bacterial colonization of different sites or are used too long for prophylaxis. The surveillance and analysis of bacteria causing serious infections in defined subsets of patients are essential for interventions aimed at improving the treatment and prophylaxis of infections in these patients. In our case, we will try to reduce the rate of multidrug-resistant GN bacteria by reducing the duration of surgical prophylaxis to 24 to 48 hours and by switching to a narrower spectrum antibiotic. The reinforcement of contact isolation and standard precautions is a process currently underway in our hospital. Mrzljak Anna M.D.*, Peric Zinaida M.D.*, Kovacevic Visnja M.D. , Gustin Denis M.D. , Vrhovac Radovan M.D.*, Tambic Andrasevic Arjana M.D. , * Departments of Medicine, University Hospital Merkur, Zagreb, Croatia, Anesthesiology and Intensive Care, University Hospital Merkur, Zagreb, Croatia, Department of Microbiology, Dr. Fran Mihaljevic University Hospital for Infectious Diseases, Zagreb, Croatia

Diabetes mellitus is a well known risk factor for infections, which remain a major cause of morbidity and mortality in patients treated with autologous peripheral blood stem cell transplantation (PBSCT). We have evaluated infectious complications following transplantation in 132 consecutive patients with relapsed or refractory Non-Hodgkin’ s lymphoma (n=101) and Hodgkin’ s disease (n=31) treated with PBSCT. Febrile neutropenia occurred in 86 (65.6%) patients at a mean of 6 days after transplantation (range 1-9, SD 1.59). Patients with diabetes mellitus had a clear predisposition for infectious complications following PBSCT. All 10 patients with diabetes developed febrile neutropenia. Compared to patients not having diabetes mellitus, they also had significantly longer median time to defervescence (4 days vs. 2 days, Logrank p=0.03). Furthermore, diabetic patients had a higher incidence of serious infections: 60% of febrile episodes in these patients (vs. 30.3% in non- diabetics) were microbiologically proven bacteremias (p=0.07). Gram positive microorganisms were responsible for the majority of documented infections with Staphylococcus epidermidis being the most frequently isolated pathogen. Infections are serious but manageable complications of PBSCT, even in patients with unfavorable risk factors. Patients with lymphoma and diabetes mellitus undergoing stem cell transplantation are especially prone to infections during the neutropenic period following transplantation. Early empirical antimicrobial therapy, tailored according to local microbiological epidemiology is essential for their optimal treatment.

Background: Infections remain a major cause of morbidity and mortality in patients with lymphoma treated with autologous hematopoietic stem cell transplantation. Knowledge of local microbiological epidemiology and patients’ risk factors are essential for optimal management of these infections. Aims: to determine incidence of febrile neutropenia (FN) in patients with lymphoma treated with autologous peripheral blood stem cell transplantation (PBSCT) and to investigate correlations of a number of variables available on the day of FN onset with the severity of infection. Patients and Methods: 102 consecutive patients (median age 44, range 19–68 yrs, 58M/44F) with relapsed or refractory Non-Hodgkin’s Lymphoma (NHL, n=78) and Hodgkin’s Disease (HD, n=24) treated with PBSCT in a single center have been evaluated for infectious complications following transplantation. Results: Febrile neutropenia occurred in 67 (65.7%) patients at a mean of 6 days after transplantation (range 1–9, SD 1.71). Microbiological work-up was done and empirical antibiotic therapy with piperacillin-tazobactam was initiated in all but 15 patients who received a carbapenem. Empirical therapy was modified in 31 (46.3%) patients. Vancomycin was added in 22 patients (32.8%), a sistemic antifungal in 4 (6.0%) and both in 6 (9.0%) patients. Three patiens died as a consequence of sepsis (TRM=2.9%). Twenty two patients (32.8%) had proven bacteremias while 27 (40.3%) had other microbiologically documented infections (MDIs). Gram+ microorganisms were responsible for the majority of all documented infections (63.6% of bacteremias and 54.5% of other MDIs). Correlations of a number of variables available at time of FN onset with the duration of febrile episode have been investigated. Patients with higher CRP levels at FN onset and those with earlier onset of fever following PBSCT had significantly longer (p=0.047 and p=0.0066, respectively) duration of fever. Other variables, including age, No. of previous lines of therapy, No. of stem cells reinfused, ANC and monocyte counts as well as peak temperature values at day of fever onset did not show statistically significant correlations with duration of fever. All patients with diabetes developed FN and, although not reaching statistical significance (p=0.09), its duration was longer in comparison to other patients. Conclusions: Infections are serious but manageable complications of PBSCT. Gram+ microorganisms remain the major cause of documented infections. Early empirical antimicrobial therapy, tailored according to local microbiological epidemiology is essential for their optimal treatment.

Infekcije predstavljaju vodeci uzrok morbiditeta i mortaliteta kod bolesnika lijecenih transplantacijom krvotvornih maticnih stanica. Poznavanje lokalne mikrobioloske epidemiologije i rizicnih cimbenika vezanih uz bolesnike vrlo su važni za njihovo optimalno lijecenje. CILJ: U bolesnika s limfomima lijecenih transplantacijom autolognih perifernih maticnih stanica (PMS) ispitati incidenciju febrilne neutropenije (FN) te istražiti povezanost varijabli izmjerenih na dan nastupa febriliteta s težinom infekcije. BOLESNICI I METODE: U istraživanje su ukljucena 102 konsekutivna bolesnika lijecena u KB Merkur od sijecnja 2002. do sijecnja 2008. (58m/44ž, dob 19- 68 uz medijan 44 godine, ne-Hodgkinov limfom=78, Hodgkinov limfom=24). REZULTATI: Febrilna se neutropenija pojavila kod 67 (65.7%) bolesnika, uz medijan nastupa 6 dana nakon reinfuzije PMS (raspon 1-9 dana, SD 1.71). Svi su bolesnici mikrobioloski obrađeni, nakon cega je zapoceta empirijska antibiotska terapija piperacilin- tazobaktamom, osim u 15 bolesnika koji su primili drugi antimikrobni lijek. Empirijska terapija je modificirana u 31 (46.3%) bolesnika. Vankomicin je dodan u 22 (32.8%) bolesnika, sistemna antifungalna terapija u 4 (6.0%), a oboje u 6 (9.0%) bolesnika. Tri su bolesnika umrla u sepsi tijekom razdoblja pancitopenije (TRM=2.9%). U 22 je bolesnika (32.8%) uzrocnik izoliran u hemokulturi, a kod 27 (40.3%) (i) u nekom drugom bioloskom uzorku. Gram+ mikroorganizmi bili su odgovorni za 63.6% svih bakterijemija, Gram- za njih 31.8%, dok je gljiva izolirana u hemokulturi 1 bolesnika (4.5% bakterijemija). Bolesnici s visim vrijednostima CRP-a i oni kod kojih je do pojave FN doslo ranije nakon transplantacije autolognih PMS imali su statisticki znacajno dulje trajanje febriliteta (p=0.047 i p=0.0066). Ostale varijable, ukljucujuci dob, broj prethodno primijenjenih linija terapije, broj reinfundiranih maticnih stanica, broj neutrofila, monocita te visina febriliteta na dan pocetka febrilne epizode nisu pokazali statisticki znacajnu korelaciju s trajanjem febriliteta. Svi su bolesnici s dijabetesom imali infektivne komplikacije, a njihovo je trajanje bilo dulje nego kod drugih bolesnika, iako ta razlika nije dostigla razinu statisticke znacajnosti (p=0.09). ZAKLJUCAK: Infekcije su opasne ali i ljecive komplikacije transplantacije autolognih PMS. Gram+ mikroorganizmi su najcesci uzrocnici infekcija u ovih bolesnika. Antimikrobnu terapiju valja primjeniti na vrijeme, a u izboru najprikladnijeg lijeka valja voditi racuna i o lokalnoj mikrobioloskoj epidemiologiji.

Bloodstream infections (BSI) are major complications of orthotopic liver transplantation (OLT) and haematopoeitic stem cell transplantation (HSCT). Knowing characteristical timing of their occurrence enables anticipation of these infections and their earlier detection, and knowing the pattern of causative microorganisms is a prerequisite for determining suitable empirical therapy. Objective: to evaluate and compare incidence, timing and aetiology of BSI post transplantation (TX) in two groups (OLT and HSCT) of patients in a single institution. Methods: 263 consecutive TXs performed from Jan 2005 to Oct 2008 have been evaluated. OLT patients: n = 136 ; mean age 49 (range 16– 75, SD 12) ; HSCT patients: n = 126 ; mean age 45 (range 19– 70, SD 14) ; autologous TX 80.2%, allogeneic 19.8%. Patients were followed up 1 year after TX ; blood culture dates and isolates were recorded. Results: 78 BSI were identified (OLT: n = 42, HSCT: n = 36) at a median of 18 (range 1– 256, SD 75) days post TX (OLT: median 42, range 1– 248 SD 65 days ; HSCT: median 8, range 1– 256, SD 85 days, Mann Whitney p = 0.004). In both groups, the majority of BSI were observed in the first trimester following TX, in HSCT patients mostly (69.4% of all BSI in HSCT) during the first two weeks (Fig. 1). Gram-negative (GN) organisms were the prevalent cause (52.6%) of BSI in both groups, with Pseudomonas aeruginosa accounting for 16.7% of all BSI and 31.7% of all GN BSI. Gram-positive (GP) pathogens were responsible for 37.2% of BSI, with coagulase-negative staphylococci being the most prevalent in this group (17.9% of all pathogens, 48.3% of all GP organisms). Mixed BSI represented 3.8% and fungaemias (candidaemias) 6.4% of all BSI. No statistically significant differences in aetiology of BSI (GN, GP, fungal) were found between OLT and HSCT patients, neither during the whole period of observation (1 year), nor in any of the 4 trimesters. A trend towards higher incidence of fungaemia in OLT and allogeneic HSCT compared to autologous HSCT patients was revealed (Chi square, p = 0.09). Conclusions: BSI are frequent complications of both OLT and HSCT, especially during the first trimester post TX. In HSCT patients they occur significantly earlier than in OLT patients. Significant differences in aetiology of BSI between OLT and HSCT patients were not found, most BSI were caused by GN organisms in both groups. Empirical therapy in these immunosuppressed patients should include agents with strong antipseudomonal activity. Figure 1. Incidence of bloodstream infections in patients after haematopoietic stem cell transplantation (HSCT) or orthotopic liver transplantation (OLT).

Background: Knowledge of local microbiological epidemiology is essential for optimal management of infections following hematopoietic stem cell transplantation (HSCT). Aims: To evaluate incidence, timing and etiology of bloodstream infections (BSI) in patients treated with HSCT in a single institution. Patients & methods: 126 consecutive transplantations (Tx) were evaluated ; 101 autologous (80.2%) and 25 allogeneic (19.8%). Results: 36 BSI were identified at a median of 8 (range 1-256, SD 85) days post Tx. The majority of BSI were observed in the first month following HSCT, mostly (69.4% of all BSI) during the first 2 weeks. A total of 12 different bacterial species were identified. Gram- organisms were isolated in 52.8% of BSI, with P.aeruginosa accounting for 25% of all BSI and 45% of all Gram- BSI. Altogether P.aeruginosa was isolated in 9 patients and 4 of these isolates were resistant to carbapenems and sensitive to only one or two other antibiotic classes. Gram+ pathogens were responsible for 36.1% of BSI, with S.epidermidis being the most prevalent in this group (22.2% of all BSI, 61.5% of all Gram+ BSI). Fungemias represented 2.8% of all BSI, a trend towards higher incidence of fungemia in allogeneic HSCT patients was observed (x2, p=0.08). Conclusions: BSI were a frequent complication of HSCT, with highest incidence during chemotherapy induced mucosal damage and neutropenia. Despite fluoroquinolone prophylaxis, more Gram- than Gram+ BSI were identified. Empirical therapy in these immunosuppressed patients should inevitably include agents with strong antipseudomonal activity ; treatment of BSI caused by P.aeruginosa remains a challenge due to the multiple resistance of this pathogen in our institution.