The Serratia marcescens Siderophore Serratiochelin Is Necessary for Full Virulence during Bloodstream Infection.

Serratia marcescens is a bacterium frequently found in the environment, but over the last several decades it has evolved into a concerning clinical pathogen, causing fatal bacteremia. To establish such infections, pathogens require specific nutrients; one very limited but essential nutrient is iron. We sought to characterize the iron acquisition systems in S. marcescens isolate UMH9, which was recovered from a clinical bloodstream infection. Using RNA sequencing (RNA-seq), we identified two predicted siderophore gene clusters (cbs and sch) that were regulated by iron. Mutants were constructed to delete each iron acquisition locus individually and in conjunction, generating both single and double mutants for the putative siderophore systems. Mutants lacking the sch gene cluster lost their iron-chelating ability as quantified by the chrome azurol S (CAS) assay, whereas the cbs mutant retained wild-type activity. Mass spectrometry-based analysis identified the chelating siderophore to be serratiochelin, a siderophore previously identified in Serratia plymuthica Serratiochelin-producing mutants also displayed a decreased growth rate under iron-limited conditions created by dipyridyl added to LB medium. Additionally, mutants lacking serratiochelin were significantly outcompeted during cochallenge with wild-type UMH9 in the kidneys and spleen after inoculation via the tail vein in a bacteremia mouse model. This result was further confirmed by an independent challenge, suggesting that serratiochelin is required for full S. marcescens pathogenesis in the bloodstream. Nine other clinical isolates have at least 90% protein identity to the UMH9 serratiochelin system; therefore, our results are broadly applicable to emerging clinical isolates of S. marcescens causing bacteremia.

Whole-genome-based analysis reveals multiclone Serratia marcescens outbreaks in a non-Neonatal Intensive Care Unit setting in a tertiary care hospital in India.

We report the use of next generation sequencing (NGS) for investigating an outbreak of 13 cases of Serratia marcescens blood stream infections in a non-Neonatal Intensive Care Unit (non-NICU) setting in a tertiary care hospital in India over 5 months. Thirteen cases of sepsis due to S. marcescens were identified in various Intensive Care Units (ICUs) over 5 months. Environmental surveillance identified isolates in the adult ICU (AICU). Antibiogram did not correlate with timeline. Sequencing libraries were prepared using Nextera XT chemistry (Illumina). Based on NGS, two clusters were identified. Cluster 1 had environmental and clinical isolates from the AICU and cluster 2 were isolates from the Coronary Care Unit (CCU). NICU and Paediatric ICU isolates did not belong to any cluster. Polyclonal outbreaks best identified by NGS can occur simultaneously. Good infection prevention practices like hand hygiene for compounded medicines and surface cleaning helped end the outbreak.

Late antibody-mediated rejection after ABO-incompatible kidney transplantation during Gram-negative sepsis.

BACKGROUND: The major challenge in ABO-incompatible transplantation is to minimize antibody-mediated rejection. Effective reduction of the anti-ABO blood group antibodies at the time of transplantation has made ABO-incompatible kidney transplantation a growing practice in our hospital and in centers worldwide. ABO antibodies result from contact with A- and B-like antigens in the intestines via nutrients and bacteria. We demonstrate a patient with fulminant antibody-mediated rejection late after ABO-incompatible kidney transplantation, whose anti-A antibody titers rose dramatically following Serratia marcescens sepsis. CASE PRESENTATION: A 58-year-old woman underwent an ABO-incompatible kidney transplantation for end-stage renal disease secondary to autosomal dominant polycystic kidney disease. It concerned a blood group A1 to O donation. Pre-desensitization titers were 64 for anti-blood group A IgM and 32 for anti-blood group A IgG titers. Desensitization treatment consisted of rituximab, tacrolimus, mycophenolate mofetil, corticosteroids, immunoadsorption and intravenous immunoglobulins. She was readmitted to our hospital 11 weeks after transplantation for S. marcescens urosepsis. Her anti-A IgM titer rose to >5000 and she developed a fulminant antibody-mediated rejection.We hypothesized that the (overwhelming) presence in the blood of S. marcescens stimulated anti-A antibody formation, as S. marcescens might share epitopes with blood group A antigen. Unfortunately we could not demonstrate interaction between blood group A and S. marcescens in incubation experiments. CONCLUSION: Two features of this post-transplant course are remarkably different from other reports of acute rejection in ABO-incompatible kidney transplantation: first, the late occurrence 12 weeks after kidney transplantation and second, the very high anti-A IgM titers (>5000), suggesting recent boosting of anti-A antibody formation by S. marcescens.

Serratia marcescens strains implicated in adverse transfusion reactions form biofilms in platelet concentrates and demonstrate reduced detection by automated culture.

BACKGROUND AND OBJECTIVES: Serratia marcescens is a gram-negative bacterium that has been implicated in adverse transfusion reactions associated with contaminated platelet concentrates. The aim of this study was to investigate whether the ability of S. marcescens to form surface-attached aggregates (biofilms) could account for contaminated platelet units being missed during screening by the BacT/ALERT automated culture system. MATERIALS AND METHODS: Seven S. marcescens strains, including biofilm-positive and biofilm-negative control strains and five isolates recovered from contaminated platelet concentrates, were grown in enriched Luria-Bertani medium and in platelets. Biofilm formation was examined by staining assay, dislodging experiments and scanning electron microscopy. Clinical strains were also analysed for their ability to evade detection by the BacT/ALERT system. RESULTS: All strains exhibited similar growth in medium and platelets. While only the biofilm-positive control strain formed biofilms in medium, this strain and three clinical isolates associated with transfusion reactions formed biofilms in platelet concentrates. The other two clinical strains, which had been captured during platelet screening by BacT/ALERT, failed to form biofilms in platelets. Biofilm-forming clinical isolates were approximately three times (P<0·05) more likely to be missed by BacT/ALERT screening than biofilm-negative strains. CONCLUSION: S. marcescens strains associated with transfusion reactions form biofilms under platelet storage conditions, and initial biofilm formation correlates with missed detection of contaminated platelet concentrates by the BacT/ALERT system.

Bacterial growth kinetics in ACD-A apheresis platelets: comparison of plasma and PAS III storage.

BACKGROUND: Our objective was to determine the growth kinetics of bacteria in leukoreduced apheresis platelets (LR-AP) in a platelet (PLT) additive solution (PAS; InterSol, Fenwal, Inc.) compared to LR-AP stored in plasma. STUDY DESIGN AND METHODS: Hyperconcentrated, double-dose LR-AP were collected from healthy donors with a separator (AMICUS, Fenwal, Inc.). LR-AP were evenly divided, InterSol was added to half (65% InterSol:35% plasma [PAS]), and PLTs in autologous plasma were used for a paired control (PL). Bacteria were inoculated into each LR-AP PAS/PL pair (0.5-1.6 colony-forming units [CFUs]/mL), and bacterial growth was followed for up to 7 days. Time to the end of the lag phase, doubling times, maximum concentration (conc-max), and time to maximum concentration (time-max) were estimated. RESULTS: Streptococcus viridans did not grow to detectable levels in either PAS or PL units. The other bacteria had no significant overall difference in the conc-max (p = 0.47) or time-max (p = 0.7) between PL and PAS LR-AP; PL had a 0.14 hours faster doubling rate (p = 0.023); and PAS had a 4.7 hours shorter lag time (p = 0.016). CONCLUSION: We observed that five index organisms will grow in LR-AP stored in a 35%:65% ratio of plasma to InterSol where initial bacterial concentrations are 0.5 to 1.6 CFUs/mL. The more rapid initiation of log-phase growth for bacteria within a PAS storage environment resulted in a bacterial concentration up to 4 logs higher in the PAS units compared to the plasma units at 24 hours, but with no difference in the conc-max. This may present an early bacterial detection advantage for PAS-stored PLTs.

Clinical experiences of the infections caused by extended-spectrum beta-lactamase-producing Serratia marcescens at a medical center in Taiwan.

CTX-M-3 has become the most common extended-spectrum beta-lactamase (ESBL) produced by Serratia marcescens in Taiwan. An expanded effort to detect ESBL among 123 nonrepetitive isolates of S. marcescens was made and 15 (12%) ESBL-producers were identified, all revealing CTX-M-3. Without routinely detecting the ESBL for S. marcescens in clinical laboratories, 80% of the ESBL-producers were reported to be susceptible to cefepime. The clinical spectrum of ESBL-producing S. marcescens-related infections included febrile urinary tract infection (n = 3); afebrile pyuria (n = 2); pneumonia (n = 3); spontaneous bacterial peritonitis (n = 3); secondary bacteremia (n = 2) and one each with primary bacteremia and colonization of the central catheter tip. Overall, the 30-day mortality rate was 33.3% (5/15) and the outcome depended on the severity of the underlying disorder and infection per se. In conclusion, although our case numbers were limited, due to the substantial incidence and associated mortality of ESBL-producing S. marcescens and its potential treatment failure by an apparently susceptible cephalosporin, we recommend that the detection and report of ESBL production for S. marcescens in clinical laboratories be made mandatory.

Serratia bacteremia in a large university hospital: trends in antibiotic resistance during 10 years and implications for antibiotic use.

OBJECTIVE: To identify antibiotic resistance trends and risk factors for resistance of Serratia species to third-generation cephalosporins. DESIGN: Retrospective survey of medical records. SETTING: A 2,200-bed, tertiary-care hospital. PATIENTS: One hundred twenty-two patients with Serratia bacteremia between January 1991 and June 2001. METHODS: Infectious disease physicians collected data from medical records regarding patient demographics, underlying disease or condition, portal of entry, microorganism, antibiogram, complications, antibiotics received, and outcome. RESULTS: Among 122 Serratia isolates, 117 (95.9%) were Serratia marcescens and 110 (90.2%) were of nosocomial origin. During the study period, the 122 isolates showed a high rate of resistance to third-generation cephalosporins (45.9%) and extended-spectrum penicillins (56.6%). The resistance rate to ciprofloxacin was 32.0%. The resistance rate to third-generation cephalosporins increased from 31.7% for 1991 to 1995 to 54.9% for 1996 to 1998 and 50.0% for 1999 to 2001. In the multivariate analysis, prior use of a second-generation cephalosporin (adjusted odds ratio [OR], 5.90; 95% confidence interval [CI90], 1.41 to 24.6; P = .015) or a third-generation cephalosporin (OR, 3.26; CI95, 1.20 to 8.87; P = .020) was a strong independent risk factor for resistance to third-generation cephalosporins. The overall case-fatality rate was 25.4% (Serratia bacteremia-related case-fatality rate, 13.1%). CONCLUSION: Prior use of a second- or third-generation cephalosporin was the most important risk factor for bacteremia with Serratia resistant to third-generation cephalosporins, suggesting the need for antibiotic control. The potential role of patient-to-patient spread could not be fully evaluated in this retrospective study.

Transfusion-related sepsis due to Serratia liquefaciens in the United States.

BACKGROUND: Severe, often fatal, transfusion reactions due to bacterial contamination of blood components continue to occur. Serratia liquefaciens, an unusual human pathogen, is a recently recognized potential cause of transfusion-related sepsis. CASE REPORTS: Five episodes of transfusion-related sepsis and endotoxic shock due to S. liquefaciens were reported to the CDC from July 1992 through January 1999. One episode has been described. The remaining four, all fatal, are described here: three associated with RBC transfusion and one associated with transfusion of platelets. In each instance, the source of contamination could not be found. The implicated units tended to be older (mean RBC age 28 days), and visual discoloration was noted in each RBC unit, although usually in retrospect. CONCLUSION: S. liquefaciens is an increasingly recognized cause of transfusion-related sepsis and is associated with a high mortality rate. S. liquefaciens can contaminate both RBCs and platelets, but the mechanism(s) of contamination remain unknown. Increased attention to pretransfusion visual inspection may avert the transfusion of some S. liquefaciens-contaminated RBC units. However, more sensitive rapid diagnostic tests are needed to further reduce the risk of transfusion-related sepsis and endotoxic shock.

Serratia type pore forming toxins.

The Serratia marcescens hemolysin represents a new type of hemolysin and has been studied in great molecular detail with regard to structure, activation and secretion. It has nothing in common with the pore forming toxins of E. coli type (RTX toxins), the Staphylococcus aureus alpha-toxin or the thiol activated toxin of group A beta-hemolytic streptococci (Streptolysin O). Studies on erythrocytes, eukaryotic cells and artificial black lipid membranes, have shown that the mechanism of pore formation of ShlA is different form other pore forming toxins. The S. marcescens hemolysin proteins ShlB and ShlA exhibit protein sequence homologues in Proteus mirabilis, Haemophilus ducreyi, Edwardsiella tarda and Erwinia chrysantemi. Furthermore, sequence motifs present in ShlA and Shlb have been shown to be important for activity and secretion of the S. marcescens hemolysin. Thus, the S. marcescens hemolysin forms the prototype of a new class of hemolysins and of a new secretory mechanism. The uniqueness of this new mechanism is underlined by the fact that activation of ShlA by ShlB strictly requires phosphatidylethanolamine as a cofactor. New data implicate a conformational change in ShlA during activation. In addition, ShlA not only forms pores in erythrocytes but also in fibroblasts and epithelial cells. The cytotoxic action of ShlA is mainly determined by lysis of infected cells in vitro. In sublytic doses, as will normally be the situation in vivo, ShlA exerts additionally effects which are currently under investigation. The knowledge of the structure, activation, secretion and mode of action of S. marcescens hemolysin has implications for proteins, related in sequence or in mode of secretion and activation.

Differential distributions in tissues and efficacies of aztreonam and ceftazidime and in vivo bacterial morphological changes following treatment.

The differential tissue distributions of aztreonam and ceftazidime within fibrin clots infected with Pseudomonas aeruginosa, Enterobacter cloacae, and Serratia marcescens, their efficacies, and the in vivo bacterial morphological changes induced by these drugs were evaluated. Rabbits were given intravenously a single dose of 100 mg of either agents/kg of body weight. In the cores of the clots, the peak levels of both drugs were much lower than those observed in the peripheries and in serum. Aztreonam's half-lives within the peripheries and in the cores of the fibrin clots were up to six times higher than observed in serum, while ceftazidime's half-lives in clots were twice that observed in serum. This resulted in a much greater penetration ratio for aztreonam than for ceftazidime. Both drugs controlled the growth of P. aeruginosa in vivo, but E. cloacae and S. marcescens responded better to ceftazidime. Morphological changes were more abundant in the peripheries than in the cores of the clots. In the control group, P. aeruginosa's morphology in the cores was different than that in the peripheries of the clots. Against P. aeruginosa, aztreonam did induce morphological changes in the cores while ceftazidime did not. Electron microscopic studies revealed that morphological changes associated with aztreonam seemed different than those of ceftazidime. Along with elongation of bacteria, more bow tie and herniated bacteria were observed with aztreonam. Though both agents selectively affect PBP 3, as manifested by elongated bacteria, they induce in the peripheries of the clots thickening, breaks, and detachment in bacterial cell walls, alterations which are generally associated with antibiotics affecting PBP 1a and 1b.

In vivo antibacterial effects of simulated human serum profiles of once-daily versus thrice-daily dosing of amikacin in a Serratia marcescens endocarditis experimental model.

Once-daily dosage of aminoglycosides is currently under consideration. The lower toxicity of this regimen has been clearly established, but there are conflicting experimental and clinical data concerning its efficacy. It is inadvisable to optimize human therapy by extrapolation from experimental studies since animal and human pharmacokinetics differ. The simulation of human pharmacokinetics in experimental infectious models would seem to offer a more rational approach. We used computer-controlled infusion of amikacin at a variable flow rate to simulate human pharmacokinetics in a Serratia marcescens rabbit endocarditis model and to compare two therapeutic regimens (once-daily versus thrice-daily doses). The doses corresponded to simulations of 15 and 30 mg/kg of body weight per day in humans, and antibacterial activity was measured in vegetations (Veg) after 24 h of treatment. The results show that the dose corresponding to 15 mg/kg/day failed to produce a significant reduction of CFU (6.8 +/- 0.9 and 6.4 +/- 0.8 log10 CFU/g of Veg, respectively, for once-daily and thrice-daily doses versus 7.6 +/- 1.0 for controls). A significant reduction was observed only for the dose corresponding to 30 mg/kg/day in humans (5.2 +/- 1.5 and 5.4 +/- 1.1 log10 CFU/g of Veg, respectively, for the two regimens). With this model, the efficacy of amikacin was similar for both regimens after 24 h of treatment simulating human pharmacokinetics.

[Critical serum concentration of antibiotics. A therapeutic tool and means of comparative evaluation]. / La concentration sérique critique des antibiotiques. Outil thérapeutique et moyen d'évaluation comparative.

It is difficult to predict the clinical activity of antibiotics solely on the basis of in vitro data. Experimental models measuring the relationship between serum concentration and in vivo activity are essential for comparing the activity of different compounds currently available. The critical serum concentration can be used to compare the intrinsic activity of antibiotics on a given bacterial strain. When compared with the minimal inhibiting concentration measured in vitro, "activity loss" can be determined for each antibiotic placed in contact with bacteria in an infected tissue. The relevance of this therapeutic tool in comparison with other methods is discussed.

Nosocomial epidemic of Serratia marcescens septicemia ascribed to contaminated blood transfusion bags.

Two cases of transfusion-related Serratia marcescens bacteremia prompted extensive epidemiologic investigations in three independent hospitals. Test tubes and plasma from donors whose blood was drawn into bags from a single production batch were cultured. Analysis of the ribotype of S. marcescens isolates was performed. For comparison, a strain from the production plant and eight other, unrelated bacteremia isolates were examined. In addition, a retrospective national survey was carried out. S. marcescens was cultured from 11 (0.73%) of 1515 blood units, and an additional (third) bacteremic patient was identified. The clinical isolates from three patients, the three units of blood transfused, and the plant-derived strain shared a unique ribotype. The incident is interpreted as a sporadic, bacterial contamination of blood bags with the S. marcescens epidemic strain, occurring during the manufacturing or packaging. A similar incident has not previously been reported. Attention is drawn to the possibility of significant contamination during the complex production of multiple-bag blood collection systems. Guidelines for improved registration and handling of transfusion complications in wards are suggested. Manufacturers should be encouraged to provide blood packs with sterile exteriors, in appropriate, single, outer packages.