Intraseasonal variation of phycocyanin concentrations and environmental covariates in two agricultural irrigation ponds in Maryland, USA.

Recently, cyanobacteria blooms have become a concern for agricultural irrigation water quality. Numerous studies have shown that cyanotoxins from these harmful algal blooms (HABs) can be transported to and assimilated into crops when present in irrigation waters. Phycocyanin is a pigment known only to occur in cyanobacteria and is often used to indicate cyanobacteria presence in waters. The objective of this work was to identify the most influential environmental covariates affecting the phycocyanin concentrations in agricultural irrigation ponds that experience cyanobacteria blooms of the potentially toxigenic species Microcystis and Aphanizomenon using machine learning methodology. The study was performed at two agricultural irrigation ponds over a 5-month period in the summer of 2018. Phycocyanin concentrations, along with sensor-based and fluorometer-based water quality parameters including turbidity (NTU), pH, dissolved oxygen (DO), fluorescent dissolved organic matter (fDOM), conductivity, chlorophyll, color dissolved organic matter (CDOM), and extracted chlorophyll were measured. Regression tree analyses were used to determine the most influential water quality parameters on phycocyanin concentrations. Nearshore sampling locations had higher phycocyanin concentrations than interior sampling locations and "zones" of consistently higher concentrations of phycocyanin were found in both ponds. The regression tree analyses indicated extracted chlorophyll, CDOM, and NTU were the three most influential parameters on phycocyanin concentrations. This study indicates that sensor-based and fluorometer-based water quality parameters could be useful to identify spatial patterns of phycocyanin concentrations and therefore, cyanobacteria blooms, in agricultural irrigation ponds and potentially other water bodies.

Antibiotic susceptibility of Legionella pneumophila strains isolated in England and Wales 2007-17.

Objectives: Antibiotic susceptibility of Legionella pneumophila is poorly understood, with treatment of Legionnaires' disease often based on empirical choice. The aim of this study was to determine the antibiotic susceptibility of L. pneumophila strains. Methods: Antibiotic susceptibility of 92 L. pneumophila strains isolated in England and Wales between 2007 and 2017 was determined using a microbroth dilution methodology for each agent tested. MICs and MBCs were determined and compared with published intracellular concentrations of each agent tested. Results: The MIC range of erythromycin was 0.06-1 mg/L, the MIC range of rifampicin was 0.0001 mg/L, the MIC range of ciprofloxacin was 0.004-0.25 mg/L and the MIC range of levofloxacin and moxifloxacin was 0.03-0.25 mg/L. The MBC range of erythromycin was 1-32 mg/L, but the MBC range of ciprofloxacin was the same as the MIC range. For levofloxacin and moxifloxacin the MBC range was elevated by one dilution and two dilutions, respectively. Typically, intracellular bronchial secretion concentrations of erythromycin might be expected to reach a suitable level to exceed the MIC range; however, 91 of 92 (98.9%) isolates had an MBC below the expected intracellular concentrations, which indicated erythromycin may have variable efficacy. MIC and MBC values of ciprofloxacin, levofloxacin and moxifloxacin were below achievable intracellular levels within bronchial secretions. Comparison of the MIC/MBC correlation showed very little clustering for erythromycin, but strong clustering for levofloxacin and to a lesser extent ciprofloxacin. Conclusions: Use of the MIC/MBC linkage analysis seems an appropriate way forward for antimicrobial susceptibility testing and supports current guidance recommending levofloxacin for the treatment of Legionnaires' disease.

The use of high-throughput sequencing to investigate an outbreak of glycopeptide-resistant Enterococcus faecium with a novel quinupristin-dalfopristin resistance mechanism.

High-throughput sequencing (HTS) has successfully identified novel resistance genes in enterococci and determined clonal relatedness in outbreak analysis. We report the use of HTS to investigate two concurrent outbreaks of glycopeptide-resistant Enterococcus faecium (GRE) with an uncharacterised resistance mechanism to quinupristin-dalfopristin (QD). Seven QD-resistant and five QD-susceptible GRE isolates from a two-centre outbreak were studied. HTS was performed to identify genes or predicted proteins that were associated with the QD-resistant phenotype. MLST and SNP typing on HTS data was used to determine clonal relatedness. Comparative genomic analysis confirmed this GRE outbreak involved two distinct clones (ST80 and ST192). HTS confirmed the absence of known QD resistance genes, suggesting a novel mechanism was conferring resistance. Genomic analysis identified two significant genetic determinants with explanatory power for the high level of QD resistance in the ST80 QD-resistant clone: an additional 56aa leader sequence at the N-terminus of the lsaE gene and a transposon containing seven genes encoding proteins with possible drug or drug-target modification activities. However, HTS was unable to conclusively determine the QD resistance mechanism and did not reveal any genetic basis for QD resistance in the ST192 clone. This study highlights the usefulness of HTS in deciphering the degree of relatedness in two concurrent GRE outbreaks. Although HTS was able to reveal some genetic candidates for uncharacterised QD resistance, this study demonstrates the limitations of HTS as a tool for identifying putative determinants of resistance to QD.

Detection and molecular characterization of Livestock-Associated MRSA in raw meat on retail sale in North West England.

Limited data are available on the prevalence of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) in the UK. We tested 124 raw meat samples for MRSA including pork (n = 63), chicken (n = 50) and turkey (n = 11) collected from retail outlets in North West England between March and July 2015. MRSA was recovered from nine (7·3%) samples (four chicken, three pork and two turkey) from different butchers and supermarkets. Four were labelled of UK origin, three were from continental Europe; the origin was not specified for two samples. Whole-genome sequencing (WGS), spa typing and the presence of lineage-specific canonical single nucleotide polymorphisms confirmed that they belonged to the livestock-associated clade of clonal complex (CC) 398. Seven (77·8%) isolates were multi-drug resistant. Phylogenetic analyses showed the isolates were diverse, suggesting multiple silent introductions of LA-MRSA into the UK food chain. Two chicken meat isolates belonged to a sub-clade recently reported from human cases in Europe where poultry meat was the probable source. The low levels of MRSA identified (<20 CFU per g) and absence of enterotoxin genes suggest the risk of acquisition of, or food-poisoning due to, LA-MRSA is low. Nevertheless, the MRSA contamination rate is higher than previously estimated; further evaluation of the public health impacts of LA-MRSA is warranted. SIGNIFICANCE AND IMPACT OF THE STUDY: Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) is a public health concern worldwide, but has only been reported sporadically in the UK. In the largest UK study to date, samples of raw meat at retail sale were examined for both the presence and levels of MRSA. We report the first isolations of CC398 LA-MRSA from poultry meat in the UK including representatives of a particular sub-clade associated with cases of human infection/colonization in Europe. Although levels were low (<20 CFU per g), the contamination rate was higher than previous UK studies. Moreover, whole-genome sequencing revealed multiple independent introductions of LA-MRSA into the UK food chain.

Depth-Dependent Survival of Escherichia coli and Enterococci in Soil after Manure Application and Simulated Rainfall.

Once released, manure-borne bacteria can enter runoff via interaction with the thin mixing layer near the soil surface. The objectives of this work were to document temporal changes in profile distributions of manure-borne Escherichia coli and enterococci in the near-surface soil layers after simulated rainfalls and to examine differences in survival of the two fecal indicator bacteria. Rainfall simulations were performed in triplicate on soil-filled boxes with grass cover and solid manure application for 1 h with rainfall depths of 30, 60, and 90 mm. Soil samples were collected weekly from depth ranges of 0 to 1, 1 to 2, 2 to 5, and 5 to 10 cm for 1 month. Rainfall intensity was found to have a significant impact on the initial concentrations of fecal indicator bacteria in the soil. While total numbers of enterococci rapidly declined over time, E. coli populations experienced initial growth with concentration increases of 4, 10, and 25 times the initial levels at rainfall treatment depths of 30, 60, and 90 mm, respectively. E. coli populations grew to the approximately the same level in all treatments. The 0- to 1-cm layer contained more indicator bacteria than the layers beneath it, and survival of indicator bacteria was better in this layer, with decimation times between 12 and 18 days after the first week of growth. The proportion of bacteria in the 0- to 1-cm layer grew with time as the total number of bacteria in the 0- to 10-cm layer declined. The results of this work indicate the need to revisit the bacterial survival patterns that are assumed in water quality models.

Distinct bacteriophages encoding Panton-Valentine leukocidin (PVL) among international methicillin-resistant Staphylococcus aureus clones harboring PVL.

Genetically diverse community-associated methicillin resistant Staphylococcus aureus (CA-MRSA) can harbor a bacteriophage encoding Panton-Valentine leukocidin (PVL) lysogenized into its chromosome (prophage). Six PVL phages (ΦPVL, Φ108PVL, ΦSLT, ΦSa2MW, ΦSa2USA, and ΦSa2958) are known, and single-nucleotide polymorphisms (SNPs) in the PVL genes have been reported. We sought to determine the distribution of lysogenized PVL phages among MRSA strains with PVL (PVL-MRSA strains), the PVL gene sequences, and the chromosomal phage insertion sites in 114 isolates comprising nine clones of PVL-MRSA that were selected for maximal underlying genetic diversity. The six PVL phages were identified by PCR; ΦSa2USA was present in the highest number of different lineages (multilocus sequence type clonal complex 1 [CC1], CC5, CC8, and sequence type 93 [ST93]) (n = 37 isolates). Analysis of 92 isolates confirmed that PVL phages inserted into the same chromosomal insertion locus in CC22, -30, and -80 but in a different locus in isolates of CC1, -5, -8, -59, and -88 and ST93 (and CC22 in two isolates). Within the two different loci, specific attachment motifs were found in all cases, although some limited inter- and intralineage sequence variation occurred. Overall, lineage-specific relationships between the PVL phage, the genes that encode the toxin, and the position at which the phage inserts into the host chromosome were identified. These analyses provide important insights into the microepidemiology of PVL-MRSA, will prove a valuable adjunct in outbreak investigation, and may help predict the emergence of new strains.

Modeling the kinetics of manure-borne fecal indicator removal in runoff.

Several manure-borne microorganism removal models have been developed to provide accurate estimations of the number of microorganisms removed from manure or manured soils undergoing rainfall. It has been commonly assumed that these models perform equally well when used to simulate microbe removal in runoff from manures of different consistency and levels of weathering. The objectives of this work were (a) to observe kinetics of the removal of Escherichia coli and enterococci with runoff for two different manure consistencies and three manure weathering durations, and (b) to compare performance of the log-linear, Vadas-Kleinman-Sharpley, and Bradford-Shijven models in simulation of the observed kinetics. Liquid and solid dairy manure were applied to grassed soil boxes that received simulated rainfall immediately after application and subsequently at 1 and 2 wk. Runoff samples were collected for 1 h at increasing time intervals during each event. Only the effective rainfall depth at the start of runoff was significantly affected by manure consistency (p = .033), whereas other parameters were not (p > .05). Substantial differences in microorganism removal kinetics during the initial, 1-, and 2-wk rainfall events were manifested by the significant (p < .05) effect of the degree of manure weathering in about 70% of cases. The log-linear model produced the largest fitting error especially during the initial rainfall event. The Vadas-Kleinman-Sharpley model and the Bradford-Schijven model were comparable in accuracy for all events. The latter model was slightly more accurate, and the former model had better expressed dependencies of parameter values on manure weathering. Ignoring manure weathering may lead to incorrect parameterization of manure removal models.

A multistep, ATP-dependent pathway for assembly of human immunodeficiency virus capsids in a cell-free system.

To understand the mechanism by which human immunodeficiency virus type 1 (HIV) capsids are formed, we have reconstituted the assembly of immature HIV capsids de novo in a cell-free system. Capsid authenticity is established by multiple biochemical and morphologic criteria. Known features of the assembly process are closely reproduced, indicating the fidelity of the cell-free reaction. Assembly is separated into co- and posttranslational phases, and three independent posttranslational requirements are demonstrated: (a) ATP, (b) a detergent-sensitive host factor, and (c) a detergent-insensitive host subcellular fraction that can be depleted and reconstituted. Assembly appears to proceed by way of multiple intermediates whose conversion to completed capsids can be blocked by either ATP depletion or treatment with nondenaturing detergent. Specific subsets of these intermediates accumulate upon expression of various assembly-defective Gag mutants in the cell-free system, suggesting that each mutant is blocked at a particular step in assembly. Furthermore, the accumulation of complexes of similar sizes in cells expressing the corresponding mutants suggests that comparable intermediates may exist in vivo. From these data, we propose a multi-step pathway for the biogenesis of HIV capsids, in which the assembly process can be disrupted at a number of discrete points.

Lactose synthetase: progesterone inhibition of the induction of alpha-lactalbumin.

Lactose synthesis in the mammary gland is dependent on the hormonally controlled synthesis of the two protein components of lactose synthetase, alpha-lactalbumin and a galactosyltransferase. Prolactin induces the synthesis of both proteins in mammary gland explants treated with insulin and hydrocortisone, but the induction kinetics cannot account for the asynchronous synthesis of the two proteins that are observed in vivo. Progesterone appears to take part in the control of lactose synthesis and acts to repress the formation of alpha-lactalbumin throughout pregnancy. At parturition, when the concentration of progesterone in the plasma decreases, the rate of alpha-lactalbumin synthesis increases.

Clinical and molecular epidemiology of ciprofloxacin-susceptible MRSA encoding PVL in England and Wales.

We aimed to enhance our case ascertainment of meticillin-resistant Staphylococcus aureus encoding Panton-Valentine leucocidin (PVL-MRSA), determine the patient demographic, risk factor and disease associations, and define the clonal diversity amongst isolates referred to the UK Health Protection Agency's Staphylococcus Reference Unit. PVL-MRSA collected during 2005-6 from community-based and hospitalised patients located across England and Wales were identified by polymerase chain reaction (PCR). Representative geographically and temporally unrelated isolates were characterised via toxin gene profiling, SCCmec, spa and agr typing, multilocus sequence typing (MLST) and minimum inhibitory concentration (MIC) determinations. PVL-MRSA were identified from 275 patients. Affected individuals were <1 to 95 years of age (mean 30, median 27 years). Forty-five isolates were from 18 household or community-based clusters and 23 isolates were from outbreaks in healthcare settings. Overall, 58% (n = 161) had skin and soft tissue infections and 9% (n = 25) presented with or developed more serious disease, including eight patients (3%) with necrotising pneumonia, five of whom subsequently died. PVL-MRSA were genetically diverse and harboured SCCmecIV or V(T)/VII. Representatives of MLST clonal complexes (CCs) 8, 30 and 80 were identified the most often. The 275 PVL-MRSA included internationally disseminated community-associated MRSA (CA-MRSA) strains, as well as other minor lineages, and were associated with typical risk factors and disease presentations.

Intraseasonal variation of E. coli and environmental covariates in two irrigation ponds in Maryland, USA.

The microbial quality of irrigation water is typically assessed by measuring the concentrations of E. coli in irrigation water reservoirs that are variable in space and time. E. coli concentrations are affected by water quality parameters that co-vary with E. coli concentrations and may be easily measured with currently available sensors. The objective of this work was to identify the most influential environmental covariates affecting E. coli concentrations during a three-month biweekly monitoring period within two irrigation ponds in Maryland during the summer of 2017. E. coli levels as well as sensor-based water quality parameters including turbidity, pH, dissolved oxygen, dissolved fluorescent organic matter, conductivity, and chlorophyll were measured at 23 and 34 locations in ponds 1 and 2, respectively. Regression tree analyses were used to determine the most influential water quality parameters for the prediction of E. coli levels. Correlations between E. coli and water quality covariates were not strong and were inconsistently significant. Shoreline sample locations had higher E. coli concentrations than interior pond samples and significant differences were observed when comparing these two groups. Regression trees provided fairly accurate predictions of E. coli levels based on water quality parameters with R2 values ranging from 0.70 to 0.93. Factors identified via the regression trees varied by sampling date but common leading covariates included cyanobacteria, organic matter, and turbidity. Results indicated environmental covariates, sensed either remotely or in situ, could be useful to delineate areas with different E. coli survival conditions across irrigation ponds and potentially other water bodies such as lakes, rivers, or bays.

The effect of fibrin-stabilizing factor on the subunit structure of human fibrin.

The formation of human fibrin from fibrinogen has been examined by polyacrylamide gel electrophoresis in sodium dodecyl sulfate, a method which separates a mixture of proteins on the basis of differences in molecular weight. It has been found that the plasma from a patient with a congenital deficiency of fibrin-stabilizing factor forms clots lacking the cross links among the alpha- and gammachains found in normal, cross-linked human fibrin. The addition of purified fibrin-stabilizing factor or normal plasma to the deficient plasma results in extensive cross-linking of the chains. Thus, the fibrinogen in the fibrin-stabilizing factor deficient plasma appears to be normal and forms fibrin which contains dimeric, cross-linked gamma-chains and polymeric, high molecular weight forms of alpha-chains. By the use of these electrophoretic methods, it has also been possible to develop a highly sensitive method for measuring the content of fibrin-stabilizing factor in plasma. This method depends upon the use of urea-treated fibrinogen, which is completely devoid of fibrin-stabilizing factor, but which forms the usual cross-linked subunits after conversion to fibrin by thrombin in the presence of fibrin-stabilizing factor.

Mechanism of ancrod anticoagulation. A direct proteolytic effect on fibrin.

Fibrin formed in response to ancrod, reptilase, or thrombin was reduced by beta-mercaptoethanol and examined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. It was found that ancrod progressively and totally digested the alpha-chains of fibrin monomers at sites different than plasmin; however, further digestion of fibrin monomers by either reptilase or thrombin was not observed. Highly purified ancrod did not activate fibrin-stabilizing factor (FSF); however, the reptilase preparation used in these experiments, like thrombin, activated FSF and thereby promoted cross-link formation. Fibrin, formed by clotting purified human fibrinogen with ancrod, reptilase, or thrombin for increasing periods of time in the presence of plasminogen, was incubated with urokinase and observed for complete lysis. Fibrin formed by ancrod was strikingly more vulnerable to plasmin digestion than was fibrin formed by reptilase or thrombin. The lysis times for fibrin formed for 2 hr by ancrod, reptilase, or thrombin were 18, 89, and 120 min, respectively. Evidence was also obtained that neither ancrod nor reptilase activated human plasminogen. These results indicate that fibrin formed by ancrod is not cross-linked and has significantly degraded alpha-chains: as expected, ancrod-formed fibrin is markedly susceptible to digestion by plasmin.

Increased Mortality in Diabetic Foot Ulcer Patients: The Significance of Ulcer Type.

Diabetic foot ulcer (DFU) patients have a greater than twofold increase in mortality compared with nonulcerated diabetic patients. We investigated (a) cause of death in DFU patients, (b) age at death, and (c) relationship between cause of death and ulcer type. This was an eleven-year retrospective study on DFU patients who attended King's College Hospital Foot Clinic and subsequently died. A control group of nonulcerated diabetic patients was matched for age and type of diabetes mellitus. The cause of death was identified from death certificates (DC) and postmortem (PM) examinations. There were 243 DFU patient deaths during this period. Ischaemic heart disease (IHD) was the major cause of death in 62.5% on PM compared to 45.7% on DC. Mean age at death from IHD on PM was 5 years lower in DFU patients compared to controls (68.2 ± 8.7 years versus 73.1 ± 8.0 years, P = 0.015). IHD as a cause of death at PM was significantly linked to neuropathic foot ulcers (OR 3.064, 95% CI 1.003-9.366, and P = 0.049). Conclusions. IHD is the major cause of premature mortality in DFU patients with the neuropathic foot ulcer patients being at a greater risk.

The bioavailability of mupirocin in nasal secretions in vitro.

AIM: To determine the bioavailability of mupirocin in human nasal secretions and to assess whether the contents of nasal secretions interact appreciably with this antibiotic. METHODS: The comparative bioavailability of mupirocin and chlorhexidine in nasal secretions was determined by bioassay after one, four, and eight hours of incubation with pooled secretions from three subjects. The interaction of mupirocin with nasal secretions was characterised by matrix assisted laser desorption time of flight mass spectrometry (MALDI-TOF). RESULTS: MALDI-TOF analysis showed that mupirocin was not absorbed by the main fraction of pooled nasal secretions and should remain active. In bioassay, mupirocin retained 100% of its antistaphylococcal activity in nasal secretions, whereas chlorhexidine was significantly reduced from 100 mg/litre to 1.5 mg/litre and from 1000 mg/litre to 38.5 mg/litre, irrespective of incubation time. CONCLUSIONS: The high bioavailability of mupirocin in nasal secretions results from the lack of appreciable molecular interactions.

Intra- and inter-generic plasmid-mediated spread of cephalosporin and aminoglycoside resistance amongst Klebsiella aerogenes K41 and other enterobacteria.

Klebsiella aerogenes K41, resistant to third generation cephalosporins and aminoglycosides, was isolated from clinical samples of 153 in-patients. Blood cultures accounted for 24 (15.7%) of isolates. The MIC(90) of ceftazidime for the isolates of 84 patients was >512 mg/l and was reduced to 2.0 by 4 mg/l of clavulanic acid, but only to 64 by 4 mg/l of sulbactam. Isolates of K. aerogenes K41 produced extended-spectrum beta-lactamase (ESBL) SHV-5 and TEM-1, identified by isoelectric focusing. Plasmid profiles showed that co-dissemination of cephalosporin and aminoglycoside resistance, plus ESBL production, coincided with the acquisition of a 116-kb plasmid. This plasmid was transferable in vitro from K. aerogenes K41 to other serotypes and genera of the Enterobacteriaceae.

The in-vitro activity of povidone-iodinecream against Staphylococcus aureus and its bioavailability in nasal secretions.

Due to the emergence of mupirocin-resistance in some epidemic strains of methicillin resistant Staphylococcus aureus (EMRSA) and the appearance of EMRSA with intermediate resistance to vancomycin, we evaluated the in-vitro activity of 5% povidone-iodine ('Betadine') cream as a possiblealternative to mupirocin for the elimination of nasal carriage of S. aureus. As judged by enrichment culture, povidone-iodine was bactericidal against three mupirocin-sensitive strains of S. aureus from nasal carriers, and against mupirocin-resistant and -sensitive strains of EMRSA types 3, 15 and 16, after incubation with povidone-iodine for 1.0 min at 32 degrees C. Mupirocin nasal ointment did not prevent growth after 180 min incubation. In a quantitative suspension test, 1:100 dilution of povidone-iodine cream completely eliminated an inoculum of 10(8)cfu/mL of all nine test organisms after incubation at 32 degrees C for 1.0 min, and 1:1000 dilution reduced cfu, by a factor of 10(5). After direct inoculation of the povidone-iodine cream to give 10(5)cfu/g, none of the test strains were recoverable after 30 s, giving a killing rate of approximately 10(4)cfu/s; for mupirocin nasal ointment, the maximum reduction of mupirocin-sensitive strains was ten fold after 3 h. Povidone-iodine activity was not detectable in sensitivity-testing agar, although 0.025% of povidone-iodine was detectable in a 15% nutrient strength tryptone soya agar. Using this minimal medium, the addition of nasal secretions (from any of 11 samples) reduced the activity of povidone-iodine by 80-90%, but mupirocin activity was unaffected. One millilitre of nasal secretions inactivated the equivalent of approximately 22.5 mg of povidone-iodine. These results suggest that povidone-iodine cream may have a role in the prevention of colonization and infection caused by MRSA, including mupirocin-resistant strains.

Minimal dose requirements for nasal mupirocin and its role in the control of epidemic MRSA.

Staphylococci are still a leading cause of hospital infection. The success of nasal mupirocin for the control of epidemic methicillin-resistant Staphylococcus aureus (EMRSA), the prevention of colonization of central venous cannulae, and the prevention of septicaemia in haemodialysis patients should encourage the use of minimal dose regimens to minimize the emergence of mupirocin resistance. Mupirocin applied to the anterior nares 4-times daily usually eliminates S. aureus, including EMRSA, within 48 h. Elimination is sustained for several weeks in patients and staff. We recently found that a single dose, or a regimen of 4-times daily for 2 days, eliminated nasal carriage of S. aureus within 24 h; 7 days after a single dose, 92% of the subjects were still cleared; 7 days after the 2-day course, 96% remained free of nasal S. aureus. Ward personnel who are nasal carriers of EMRSA can, provided that other carriage sites are negative, return to work after 2 days of a 4-times daily intranasal regimen. The UK guidelines, recently published in this Journal, recommend an aggressive approach to identifying and eliminating EMRSA, including the elimination of nasal carriage. This approach is increasingly associated with the control of EMRSA in the UK and elsewhere.

Reduction in the colonization of central venous cannulae by mupirocin.

In an in-vitro simulation of an intravascular cannula enclosed in a fibrin sheath, 0.03 mg1(-1) of mupirocin prevented significant colonization [greater than 15 colony forming units (cfu)] by two clinical isolates of Staphylococcus epidermidis and one each of S. saprophyticus, S. hominis and S. haemolyticus. This suggests that in vivo, where protein binding of mupirocin is 95-97%, 1 mg 1(-1) of mupirocin at the cannula surface would be required to prevent colonization. These results support the findings of our previously published prospective controlled trial, in which mupirocin applied to the insertion sites of 172 internal jugular cannulae reduced the rate of colonization of cannula tips to 5%, compared with 25% for the 186 controls (P less than 0.001). Of the 46 colonized cannula tips from 110 control patients, the same species was isolated from the skin of the insertion site in 67% and from the lumen flush in only 15%. Analysed by patient, mupirocin reduced the proportion of patients with colonized tips from 17% to 3% after 24 h of infection, and from 35 to 10% after 48 h (P = 0.002). The use of agar containing charcoal, as a mupirocin neutralizer, and the incubation of tip-culture plates flooded with the Oxford staphylococcus, gave no evidence of carry over of mupirocin onto cannulae removed from mupirocin-treated patients.

Mupirocin for the reduction of colonization of internal jugular cannulae--a randomized controlled trial.

In a prospective study, 218 cardiothoracic patients, in whom 'Abbocath-T' cannulae had been inserted preoperatively into the internal jugular vein, were randomized to receive skin preparation of the insertion site with tincture of iodine (108 controls) or tincture of iodine followed by application of sterile 2% calcium mupirocin ointment (110 test patients). Cannulae were usually removed within 48 h of the operation. Patients receiving mupirocin were less likely to develop significant colonization of one or more of their cannulae as judged by Maki's criterion of a yield of greater than 15 colony forming units (cfu) from a cannula segment rolled on an agar plate (17% of mupirocin treated patients compared with 54% of the controls, P less than 0.001). Coagulase-negative staphylococci, micrococci, or both, were the commonest isolates and were cultured from 70% of the 186 control cannulae compared with 24% of 172 cannulae inserted through mupirocin-treated skin (P less than 0.001). A count of more than 15 cfu was found on the tips of 25% control cannulae compared with 5% of the cannulae from mupirocin-treated patients, an effect which was independent of in-situ time (P less than 0.001). For cannulae with colonized tips, the same species was isolated from the skin of the insertion site in 67%, from the exterior of the hub in 61% and from the lumen in only 15%. There were no side effects attributed to mupirocin or superinfection with resistant organisms. We conclude that in cardiothoracic patients the application of mupirocin after standard skin preparation with tincture of iodine significantly reduces the colonization of central venous cannulae by organisms derived from the skin insertion site.