Whole Genome Sequencing of Invasive Neonatal Escherichia coli from Uppsala County, Sweden.

BACKGROUND: This study sought to investigate associations between a virulence factors and phylogeny in all neonatal E. coli bloodstream infections from patients admitted to the neonatal intensive care unit at Uppsala University Hospital between 2005 to 2020. METHODS: A total of 37 E. coli isolates from 32 neonates were whole genome sequenced and analysed for virulence factors related to extraintestinal E. coli, patient-related data were collected retrospectively in the medical records. RESULTS: E. coli isolates that belong to phylogroup B2 were associated with mortality (OR 26, p < 0.001), extreme prematurity with delivery before gestational week 28 (OR 9, p < 0.05) and shock (OR 9, p < 0.05) compared with isolates of non-B2 group. Female neonates were more often infected by isolates of phylogroup B2 E. coli compared with male neonates (OR 7, p = 0.05). The identification of the genotoxin determinant clb coding for colibactin exhibited strong associations with mortality (OR 67, p < 0.005), gestational age (OR 18, p < 0.005), and shock (OR 26, p < 0.005). DISCUSSION: The study highlighted the correlation between neonatal E. coli bacteraemia caused by phylogroup B2 and the role of colibactin. Moreover, it emphasised sex-based differences in bloodstream infections among the bacterial population of E. coli.

Extension of Pharmacokinetic/Pharmacodynamic Time-Kill Studies To Include Lipopolysaccharide/Endotoxin Release from Escherichia coli Exposed to Cefuroxime.

The release of inflammatory bacterial products, such as lipopolysaccharide (LPS)/endotoxin, may be increased upon the administration of antibiotics. An improved quantitative understanding of endotoxin release and its relation to antibiotic exposure and bacterial growth/killing may be gained by an integrated analysis of these processes. The aim of this work was to establish a mathematical model that relates Escherichia coli growth/killing dynamics at various cefuroxime concentrations to endotoxin release in vitro Fifty-two time-kill experiments informed bacterial and endotoxin time courses and included both static (0×, 0.5×, 1×, 2×, 10×, and 50× MIC) and dynamic (0×, 15×, and 30× MIC) cefuroxime concentrations. A model for the antibiotic-bacterium interaction was established, and antibiotic-induced bacterial killing followed a sigmoidal Emax relation to the cefuroxime concentration (MIC-specific 50% effective concentration [EC50], maximum antibiotic-induced killing rate [Emax] = 3.26 h-1 and γ = 3.37). Endotoxin release was assessed in relation to the bacterial processes of growth, antibiotic-induced bacterial killing, and natural bacterial death and found to be quantitatively related to bacterial growth (0.000292 endotoxin units [EU]/CFU) and antibiotic-induced bacterial killing (0.00636 EU/CFU). Increased release following the administration of a second cefuroxime dose was described by the formation and subsequent antibiotic-induced killing of filaments (0.295 EU/CFU). Release due to growth was instantaneous, while release due to antibiotic-induced killing was delayed (mean transit time of 7.63 h). To conclude, the in vitro release of endotoxin is related to bacterial growth and antibiotic-induced killing, with higher rates of release upon the killing of formed filaments. Endotoxin release over 24 h is lowest when antibiotic exposure rapidly eradicates bacteria, while increased release is predicted to occur when growth and antibiotic-induced killing occur simultaneously.

Antibiotic susceptibility testing in less than 30 min using direct single-cell imaging.

The emergence and spread of antibiotic-resistant bacteria are aggravated by incorrect prescription and use of antibiotics. A core problem is that there is no sufficiently fast diagnostic test to guide correct antibiotic prescription at the point of care. Here, we investigate if it is possible to develop a point-of-care susceptibility test for urinary tract infection, a disease that 100 million women suffer from annually and that exhibits widespread antibiotic resistance. We capture bacterial cells directly from samples with low bacterial counts (104 cfu/mL) using a custom-designed microfluidic chip and monitor their individual growth rates using microscopy. By averaging the growth rate response to an antibiotic over many individual cells, we can push the detection time to the biological response time of the bacteria. We find that it is possible to detect changes in growth rate in response to each of nine antibiotics that are used to treat urinary tract infections in minutes. In a test of 49 clinical uropathogenic Escherichia coli (UPEC) isolates, all were correctly classified as susceptible or resistant to ciprofloxacin in less than 10 min. The total time for antibiotic susceptibility testing, from loading of sample to diagnostic readout, is less than 30 min, which allows the development of a point-of-care test that can guide correct treatment of urinary tract infection.

Dynamics of Endotoxin, Inflammatory Variables, and Organ Dysfunction After Treatment With Antibiotics in an Escherichia coli Porcine Intensive Care Sepsis Model.

OBJECTIVES: To investigate the dynamics of antibiotic-induced endotoxin liberation and inflammatory response in vivo in a clinically relevant large animal intensive care sepsis model and whether the addition of an aminoglycoside to a ß-lactam antibiotic affects these responses. DESIGN: Prospective, placebo-controlled interventional experimental study. SETTING: University research unit. SUBJECTS: Thirty-six healthy pigs administered Escherichia coli as a 3-hour infusion. INTERVENTIONS: After 2 hours, during E. coli infusion, the animals were exposed to cefuroxime alone, the combination of cefuroxime and tobramycin, or saline. MEASUREMENTS AND MAIN RESULTS: Plasma endotoxin, interleukin-6, tumor necrosis factor-α, leucocytes, and organ dysfunction were recorded for 4 hours after antibiotic treatment, and differences to the values before treatment were calculated. In vitro experiments were performed to ascertain whether endotoxin is released during antibiotic-induced bacterial killing of this E. coli strain. Despite differences between the treatment arms in vitro, no differences in plasma endotoxin were observed in vivo. Antibiotic-treated animals demonstrated a higher interleukin-6 response (p < 0.001), greater leucocyte activation (p < 0.001), and more pronounced deterioration in pulmonary static compliance (p < 0.01) over time than controls. Animals treated with the combination showed a trend toward less inflammation. CONCLUSIONS: Treatment with antibiotics may elicit an increased inflammatory interleukin-6 response that is associated with leucocyte activation and pulmonary organ dysfunction. No observable differences were detected in plasma endotoxin concentrations. The reduction in cefuroxime-induced endotoxin release after the addition of an aminoglycoside in vitro could not be reproduced in this model.

A general method for rapid determination of antibiotic susceptibility and species in bacterial infections.

To ensure correct antibiotic treatment and reduce the unnecessary use of antibiotics, there is an urgent need for new rapid methods for species identification and determination of antibiotic susceptibility in infectious pathogenic bacteria. We have developed a general method for the rapid identification of the bacterial species causing an infection and the determination of their antibiotic susceptibility profiles. An initial short cultivation step in the absence and presence of different antibiotics was combined with sensitive species-specific padlock probe detection of the bacterial target DNA to allow a determination of growth (i.e., resistance) and no growth (i.e., susceptibility). A proof-of-concept was established for urinary tract infections in which we applied the method to determine the antibiotic susceptibility profiles of Escherichia coli for two drugs with 100% accuracy in 3.5 h. The short assay time from sample to readout enables fast appropriate treatment with effective drugs and minimizes the need to prescribe broad-spectrum antibiotics due to unknown resistance profiles of the treated infection.

Effect of glyceryl trinitrate on staphylococcus aureus growth and leukocyte activation during simulated extracorporeal circulation.

BACKGROUND: Previously, nitric oxide has been shown to possess antimicrobial effects. In this study, we aim to test the effect of glyceryl trinitrate (GTN) on Staphylococcus aureus growth during simulated extracorporeal circulation (SECC) and also to examine the effect of S. aureus, alone and in combination with GTN, on activation markers of the innate immune system during SECC. METHODS: In an in vitro system of SECC, we measured GTN-induced changes in markers of leukocyte activation in whole blood caused by S. aureus infestation, as well as the effect of GTN on S. aureus growth. RESULTS: GTN had no effect on S. aureus growth after 240 minutes SECC. Staphylococcus aureus reduced the expression of granulocyte Fcγ-receptor CD32 but stimulated the expression of monocyte CD32. Staphylococcus aureus stimulated expression of some leukocyte adhesion key proteins, activation marker CD66b, lipopolysaccharide-receptor CD14, and C3b-receptor CD35. Staphylococcus aureus and GTN addition induced significant increases in monocyte CD63 (lysosomal granule protein) levels. CONCLUSION: GTN does not affect S. aureus growth during SECC and has no effect on SECC-induced leukocyte activation.

Limitations in predicting reduced susceptibility to third generation cephalosporins in Escherichia coli based on whole genome sequence data.

Prediction of antibiotic resistance from whole genome sequence (WGS) data has been proposed. However, the performance of WGS data analysis for this matter may be influenced by the resistance mechanism's biology. This study compared traditional antimicrobial susceptibility testing with whole genome sequencing for identification of extended-spectrum beta-lactamases (ESBL) in a collection of 419 Escherichia coli isolates. BLASTn-based prediction and read mapping with srst2 gave matching results, and in 381/419 (91%) isolates WGS was congruent with phenotypic testing. Incongruent results were grouped by potential explanations into biological-related and sequence analysis-related results. Biological-related explanations included weak ESBL-enzyme activity (n = 4), inconclusive phenotypic ESBL-testing (n = 4), potential loss of plasmid during subculturing (n = 7), and other resistance mechanisms than ESBL-enzymes (n = 2). Sequence analysis-related explanations were cut-off dependency for read depth (n = 5), too stringent (n = 3) and too loose cut-off for nucleotide identity and coverage (n = 13), respectively. The results reveal limitations of both traditional antibiotic susceptibility testing and sequence-based resistance prediction and highlight the need for evidence-based standards in sequence analysis.

The First Swedish Outbreak with VIM-2-Producing Pseudomonas aeruginosa, Occurring between 2006 and 2007, Was Probably Due to Contaminated Hospital Sinks.

Multidrug-resistant Pseudomonas aeruginosa is an increasing clinical problem worldwide. The aim of this study was to describe the first outbreak of a Verona integron-borne metallo-ß-lactamase (VIM)-2-producing P. aeruginosa strain in Sweden and its expansion in the region. A cluster of multidrug-resistant P. aeruginosa appeared at two neighbouring hospitals in 2006. The isolates were characterized by PCR, pulsed-field gel electrophoresis (PFGE), and whole-genome sequencing. Patient charts, laboratory records, and hygiene routines were reviewed, and patients, staff, and the environment were screened. The investigation revealed a clonal outbreak of a VIM-2-producing P. aeruginosa strain belonging to the high-risk clonal complex 111, susceptible only to gentamicin and colistin. No direct contact between patients could be established, but most of them had stayed in certain rooms/wards weeks to months apart. Cultures from two sinks yielded growth of the same strain. The outbreak ended when control measures against the sinks were taken, but new cases occurred in a tertiary care hospital in the region. In conclusion, when facing prolonged outbreaks with this bacterium, sinks and other water sources in the hospital environment should be considered. By implementing proactive control measures to limit the bacterial load in sinks, the waterborne transmission of P. aeruginosa may be reduced.

Effects of silver-based wound dressings on the bacterial flora in chronic leg ulcers and its susceptibility in vitro to silver.

Silver-based dressings have been used extensively in wound management in recent years, but data on their antimicrobial activity in the clinical setting are limited. In order to explore their effects on chronic leg ulcer flora, 14 ulcers were cultured after at least 3 weeks treatment with Aquacel Ag(®) or Acticoat(®). Phenotypic and genetic silver resistance were investigated in a total of 56 isolates. Silver-based dressings had a limited effect on primary wound pathogens, which were present in 79% of the cultures before, and 71% after, treatment. One silver-resistant Enterobacter cloacae strain was identified (silver nitrate minimal inhibitory concentration (MIC) > 512 mg/l, positive for silE, silS and silP). Further studies in vitro showed that inducible silver-resistance was more frequent in Enterobacteriaceae with cephalosporin-resistance and that silver nitrate had mainly a bacteriostatic effect on Staphylococcus aureus. Monitoring of silver resistance should be considered in areas where silver is used extensively.

Infection control measures to stop the spread of sequence type 15 OXA-23-producing Acinetobacter baumannii in a Swedish Burn Center.

OBJECTIVE: To describe the course of the outbreak and infection control measures to stop the spread of sequence type 15 OXA-23-producing Acinetobacter baumannii in the Burn Center of Uppsala University Hospital, between November 2014 and the end of April 2015. METHODS: Compliance with hand hygiene, dress code, and cleaning routines were reviewed, the ward's environment was systematically investigated to identify potential environmental sources. Sampling routines for A. baumannii, from patients and environment, were established, and the epidemiological relationship was analysed for all carbapenem-resistant A. baumannii isolates using arbitrarily primed polymerase chain reaction (AP-PCR) and pulsed-field gel electrophoresis (PFGE). RESULTS: A total of 54 patients were treated at the burn intensive care unit during the studied, approximately five months period, and an OXA-23-producing A. baumannii was isolated from nine patients (9/54, 17%), whereof two died (2/9, 22.2%). All isolates shared identical PFGE-genotype patterns and belonged to sequence type 15; AP-PCR was eligible for prompt epidemiological investigations. CONCLUSIONS: Higher awareness and increased compliance with hand hygiene and dress code as well as intensified cleaning protocols of the environment and equipment were successfully established and likely to have led to stop the spread of sequence type 15 OXA-23-producing Acinetobacter baumannii.

Molecular Characterization of Multidrug-Resistant Yersinia enterocolitica From Foodborne Outbreaks in Sweden.

The foodborne pathogen Yersinia enterocolitica causes gastrointestinal infections worldwide. In the spring of 2019, the Swedish Public Health Agency and Statens Serum Institut in Denmark independently identified an outbreak caused by Yersinia enterocolitica 4/O:3 that after sequence comparison turned out to be a cross-border outbreak. A trace-back investigation suggested shipments of fresh prewashed spinach from Italy as a common source for the outbreak. Here, we determined the genome sequences of five Y. enterocolitica clinical isolates during the Swedish outbreak using a combination of Illumina HiSeq short-read and Nanopore Technologies' MinION long-read whole-genome sequencing. WGS results showed that all clinical strains have a fully assembled chromosome of approximately 4.6 Mbp in size and a 72-kbp virulence plasmid; one of the strains was carrying an additional 5.7-kbp plasmid, pYE-tet. All strains showed a high pathogen probability score (87.5%) with associated genes for virulence, all of which are closely related to an earlier clinical strain Y11 from Germany. In addition, we identified a chromosomally encoded multidrug-resistance cassette carrying resistance genes against chloramphenicol (catA1), streptomycin (aadA1), sulfonamides (sul1), and a mercury resistance module. This chromosomally encoded Tn2670 transposon has previously been reported associated with IncFII plasmids in Enterobacteriaceae: a Shigella flexneri clinical isolate from Japan in 1950s, a Klebsiella pneumoniae outbreak from Australia in 1997, and Salmonella enterica serovar Typhimurium. Interestingly, we identified an additional 5.7-kbp plasmid with tetB (encoding an ABC transporter), Rep, and its own ORI and ORIt sites, sharing high homology with small tetB-Rep plasmids from Pasteurellaceae. This is the first time that Tn2670 and Pasteurellaceae plasmids have been reported in Y. enterocolitica. Taken together, our study showed that the Swedish Y. enterocolitica outbreak strains acquired multi-antibiotic and metal-resistance genes through horizontal gene transfer, suggesting a potential reservoir of intraspecies dissemination of multidrug-resistance genes among foodborne pathogens. This study also highlights the concern of food-chain contamination of prewashed vegetables as a perpetual hazard against public health.

Ultraviolet-C decontamination of a hospital room: Amount of UV light needed.

INTRODUCTION: Our primary aim was to investigate, using a commercial radiometer, the ultraviolet C (UVC) dose received in different areas in a burn ICU ward room after an automated UVC decontamination. The secondary aim was to validate a disposable UVC-dose indicator with the radiometer readings. METHODS: Disposable indicators and an electronic radiometer were positioned in ten different positions in a burn ICU room. The room was decontaminated using the Tru-D™-UVC device. Colour changes of the disposable indicators and radiometer readings were noted and compared. Experiment was repeated 10 times. FINDINGS: The UVC radiation received in different areas varied between 15.9mJ/cm2 and 1068mJ/cm2 (median 266mJ/cm2). Surfaces, at shorter distances and in the direct line of sight of the UVC device showed statistically significant higher UVC doses than surfaces in the shadow of equipment (p=0.019). The UVC-dose indicator's colour change corresponded with the commercially radiometer readings. CONCLUSIONS: The amount of UVC radiation that is received in surfaces depends on their locations in the room (ie distance from the UVC emitter) and whether any objects shadow the light. In this study we suggest that quality controls should be used to assure that enough UVC radiation reaches all surfaces.

Distribution of class 1 integrons in historic and contemporary collections of human pathogenic Escherichia coli.

Integrons play a major role in the evolution and spread of antimicrobial resistance in human pathogens, including Escherichia coli. This study describes the occurrence of class 1 integrons in human pathogenic E. coli, in three isolate collections involving three periods from the last 100 years (i) the Murray collection (n = 58 bacteria isolated from the 1910s to 1940s); (ii) the E. coli reference (ECOR) collection (n = 37 isolates mainly from the 1980s); and (iii) a recently assembled collection (n = 88 isolates obtained in 2016). High-quality whole genome sequences (WGSs) were available for all isolates. Integrons were detected in the WGSs with the program IntegronFinder and the results compared with three established methods: (i) polymerase chain reaction detection of the integrase gene; (ii) BLAST searching using draft genomes; and (iii) mapping of short reads. No integrons were found in any of the Murray Collection isolates; however, integrons were present in 3% of the isolates from ECOR collection, assembled in the 1980s, and 26% of the isolates from the 2010s. Similarly, antimicrobial resistance determinants were not present in the Murray Collection isolates, whereas they were present in 19% of the ECOR Collection isolates and in 55% of the isolates obtained in during the 2010s.

Inflammatory and circulatory effects of the reduction of endotoxin concentration in established porcine endotoxemic shock--a model of endotoxin elimination.

OBJECTIVE: To study whether a reduction of the endotoxin load, once a generalized inflammatory state has been established, reduces the inflammatory response and endotoxin-induced effects on circulation, hypoperfusion, and organ dysfunction. DESIGN: Prospective parallel-grouped placebo-controlled randomized interventional experimental study. SETTING: University research unit. SUBJECTS: Healthy pigs. INTERVENTIONS: The animals were subjected to a continuous endotoxin infusion rate of either 4.0 or 0.063 microg endotoxin x kg x h for 1, 2, or 6 hours. The 1- and 2-hour infusion groups represented the applied therapy by a reduction of the endotoxin load of 5/6 and 2/3, respectively. MEASUREMENTS AND MAIN RESULTS: During a 6-hour experiment, laboratory and physiologic parameters were recorded hourly in 26 anesthetized and mechanically ventilated pigs. Primary end point was to detect differences in tumor necrosis factor-alpha (TNF-alpha) concentration during the last 3 hours of the experiment. Despite the early reduction of the endotoxin load, no effect on TNF-alpha concentration was observed. Similarly, in circulatory parameters, such as mean arterial pressure and oxygen delivery, and in platelet count and renal function, no effects were noted. However, there was some improvement in pulmonary compliance and function as determined by Pao2, Paco2, and pH. These changes were associated with slight improvements in leukocyte response and capillary leakage. CONCLUSIONS: Termination of the endotoxin infusion represents an incontestable model of endotoxin concentration reduction. Endotoxin elimination strategies applied at the TNF-alpha peak or later will have very little or no effect on TNF-alpha-mediated toxicity. Nevertheless, there was an effect on the leukocyte response that was associated with an improvement in respiratory function and microcirculation, making it impossible to rule out fully the beneficial effect of this strategy. However, the effects were limited in relation to the magnitude of the endotoxin concentration reduction and the very early application of the antiendotoxin measure.

The impact of the systemic inflammatory response on hepatic bacterial elimination in experimental abdominal sepsis.

BACKGROUND: Bacterial translocation from the gut has been suggested to induce a systemic inflammatory response syndrome (SIRS) and organ dysfunction. The liver has a pivotal role in eliminating circulating bacteria entering from the gut. We investigated whether pre-existing inflammation affects hepatic bacterial elimination. METHODS: Fifteen anaesthetised piglets were infused with E. coli in the portal vein for 3 h. The naive group (n = 6) received the bacterial infusion without endotoxin exposure. SIRS (SIRS group, n = 6) was induced by endotoxin infusion 24 h before the bacterial infusion. For effects of anaesthesia, controls (n = 3) received saline instead of endotoxin for 24 h. Bacterial counts and endotoxin levels in the portal and hepatic veins were analysed during bacterial infusion. RESULTS: The bacterial killing rate was higher in the naive group compared with the SIRS group (p = 0.001). The ratio of hepatic to portal venous bacterial counts, i.e. the median bacterial influx from the splanchnic circulation, was 0.06 (IQR 0.01-0.11) in the naive group and 0.71 (0.03-1.77) in the SIRS group at 3 h, and a magnitude lower in the naive group during bacteraemia (p = 0.03). Similar results were seen for hepatic endotoxin elimination. Peak log tumour necrosis factor alpha was higher in the naive 4.84 (4.77-4.89) vs. the SIRS group 3.27 (3.26-3.32) mg/L (p < 0.001). CONCLUSIONS: Our results suggest that hepatic bacterial and endotoxin elimination is impaired in pigs with pre-existing SIRS while the inflammatory response to bacterial infusion is diminished. If similar mechanisms operate in human critical illness, the hepatic elimination of bacteria from the gut could be impaired by SIRS.

A non-linear mixed effect model for innate immune response: In vivo kinetics of endotoxin and its induction of the cytokines tumor necrosis factor alpha and interleukin-6.

Endotoxin, a component of the outer membrane of Gram-negative bacteria, has been extensively studied as a stimulator of the innate immune response. However, the temporal aspects and exposure-response relationship of endotoxin and resulting cytokine induction and tolerance development is less well defined. The aim of this work was to establish an in silico model that simultaneously captures and connects the in vivo time-courses of endotoxin, tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), and associated tolerance development. Data from six studies of porcine endotoxemia in anesthetized piglets (n = 116) were combined and used in the analysis, with purified endotoxin (Escherichia coli O111:B4) being infused intravenously for 1-30 h in rates of 0.063-16.0 µg/kg/h across studies. All data were modelled simultaneously by means of importance sampling in the non-linear mixed effects modelling software NONMEM. The infused endotoxin followed one-compartment disposition and non-linear elimination, and stimulated the production of TNF-α to describe the rapid increase in plasma concentration. Tolerance development, observed as declining TNF-α concentration with continued infusion of endotoxin, was also driven by endotoxin as a concentration-dependent increase in the potency parameter related to TNF-α production (EC50). Production of IL-6 was stimulated by both endotoxin and TNF-α, and four consecutive transit compartments described delayed increase in plasma IL-6. A model which simultaneously account for the time-courses of endotoxin and two immune response markers, the cytokines TNF-α and IL-6, as well as the development of endotoxin tolerance, was successfully established. This model-based approach is unique in its description of the time-courses and their interrelation and may be applied within research on immune response to bacterial endotoxin, or in pre-clinical pharmaceutical research when dealing with study design or translational aspects.

The first major extended-spectrum beta-lactamase outbreak in Scandinavia was caused by clonal spread of a multiresistant Klebsiella pneumoniae producing CTX-M-15.

Between May and December 2005, 64 multidrug-resistant isolates of Klebsiella pneumoniae were detected from patients admitted to Uppsala University Hospital. This represented a dramatic increase in ESBL-producing K. pneumoniae compared to previous years. To investigate the epidemiology and to characterize the resistance mechanisms of the isolates, a study was initiated. Antibiotic susceptibility was determined by means of the Etest and the disc diffusion method. Extended-spectrum beta-lactamase (ESBL) production was identified by clavulanic acid synergy test and confirmed with PCR amplification followed by DNA sequencing. DNA profiles of the isolates were examined with pulsed-field gel electrophoresis (PFGE). All isolates were resistant or exhibited reduced susceptibility to cefadroxil, cefuroxime, cefotaxime, ceftazidime, aztreonam, piperacillin/tazobactam, ciprofloxacin, tobramycin, and trimethoprim-sulfamethoxazole. They produced ESBL of the CTX-M-15 type, and the involvement of a single K. pneumoniae clone was shown. This is the first major clonal outbreak of multiresistant ESBL-producing K. pneumoniae in Scandinavia. The outbreak demonstrates the epidemic potential of enterobacteria containing ESBLs of the CTX-M type, even in a country with a relatively low selective pressure and a low prevalence of multiresistant bacteria.

Detection of Campylobacter in human and animal field samples in Cambodia.

Campylobacter are zoonotic bacteria and a leading cause of human gastroenteritis worldwide with Campylobacter jejuni and C. coli being the most commonly detected species. The aim of this study was to detect Campylobacter in humans and livestock (chickens, ducks, pigs, cattle, water buffalo, quail, pigeons and geese) in rural households by routine culturing and multiplex PCR in faecal samples frozen before analysis. Of 681 human samples, 82 (12%) tested positive by PCR (C. jejuni in 66 samples and C. coli in 16), but none by routine culture. Children were more commonly Campylobacter positive (19%) than adult males (8%) and females (7%). Of 853 livestock samples, 106 (12%) tested positive by routine culture and 352 (41%) by PCR. Campylobacter jejuni was more frequent in chickens and ducks and C. coli in pigs. In conclusion, Campylobacter proved to be highly prevalent by PCR in children (19%), ducks (24%), chickens (56%) and pigs (72%). Routine culturing was insufficiently sensitive in detecting Campylobacter in field samples frozen before analysis. These findings suggest that PCR should be the preferred diagnostic method for detection of Campylobacter in humans and livestock where timely culture is not feasible.

Endotoxin neutralization and anti-inflammatory effects of tobramycin and ceftazidime in porcine endotoxin shock.

INTRODUCTION: Antibiotics used for treatment of severe bacterial infections have been shown to exert effects on the inflammatory response in addition to their antibacterial effects. The aim of the present study was to investigate whether the biological effects of endotoxin in a porcine model could be neutralized by tobramycin, and whether tobramycin or ceftazidime was able to modulate the inflammatory response. METHOD: Thirteen piglets were subjected to endotoxin infusion at an initial rate of 4 microgram/kg per hour, which was reduced to 1 microgram/kg per hour after 30 min. Before endotoxin infusion, the animals received saline (n = 4), ceftazidime (n = 5), or tobramycin (n = 4) at clinically relevant doses. Physiological parameters were measured and blood samples were taken hourly for 6 hours for analysis of tumour necrosis factor-alpha, IL-6 and endotoxin concentrations. RESULTS: All of the animals exhibited physiological signs of severe sepsis without major differences between the groups. Plasma endotoxin concentration was stable after 1 hour. There were no differences in endotoxin concentration or initial tumour necrosis factor-alpha and IL-6 concentrations between the groups. At 6 hours the IL-6 concentration was significantly lower in the ceftazidime group than in the saline group (P < 0.05), and in both the ceftazidime and the tobramycin groups there were significantly greater reductions from peak values (P < 0.05). CONCLUSION: There was no neutralization of the biological effects of endotoxin in this porcine model. However, our data indicate a possible anti-inflammatory effect exerted by both ceftazidime and tobramycin, which manifested as a significantly greater reduction in IL-6 in comparison with the untreated group.

Level of decontamination after washing textiles at 60°C or 70°C followed by tumble drying.

BACKGROUND: Several major outbreaks in healthcare facilities have occurred with the emergence of multi-resistant bacteria. A possible route for dissemination is the hospital textiles and inadequate laundering of them. The aim of this study was to develop an easy-to-use method for simulating the laundering process of hospital textiles, and thereafter apply the method when evaluating the decontaminating efficacy of two different washing temperatures. METHODS: The laundering process, including tumble drying, took place at two professional laundries. Enterococcus faecium was used as bioindicator. RESULTS: The results showed that a lowering of the washing temperature from 70°C to 60°C did not affect the decontamination efficacy; the washing cycle alone reduced the number of bacteria with 3-5 log10 CFU, whereas the following tumble drying reduced the bacterial numbers with another 3-4 log10 CFU, yielding the same final result independent of washing temperature. Without tumble drying, there was an obvious risk of adding non-fermenting gram-negative bacteria to the fabric. These bacteria originated from the washing cycle. CONCLUSION: A simple method to simulate hospital laundering was developed. To save energy, it is possible to use a washing temperature of 60°C, but the washing cycle should be followed by tumble drying, and the whole laundering process needs to be monitored to maintain sufficient textile hygiene.