Distinct patterns of tissue-specific lipid accumulation during the induction of insulin resistance in mice by high-fat feeding.

AIMS/HYPOTHESIS: While it is well known that diet-induced obesity causes insulin resistance, the precise mechanisms underpinning the initiation of insulin resistance are unclear. To determine factors that may cause insulin resistance, we have performed a detailed time-course study in mice fed a high-fat diet (HFD). METHODS: C57Bl/6 mice were fed chow or an HFD from 3 days to 16 weeks and glucose tolerance and tissue-specific insulin action were determined. Tissue lipid profiles were analysed by mass spectrometry and inflammatory markers were measured in adipose tissue, liver and skeletal muscle. RESULTS: Glucose intolerance developed within 3 days of the HFD and did not deteriorate further in the period to 12 weeks. Whole-body insulin resistance, measured by hyperinsulinaemic-euglycaemic clamp, was detected after 1 week of HFD and was due to hepatic insulin resistance. Adipose tissue was insulin resistant after 1 week, while skeletal muscle displayed insulin resistance at 3 weeks, coinciding with a defect in glucose disposal. Interestingly, no further deterioration in insulin sensitivity was observed in any tissue after this initial defect. Diacylglycerol content was increased in liver and muscle when insulin resistance first developed, while the onset of insulin resistance in adipose tissue was associated with increases in ceramide and sphingomyelin. Adipose tissue inflammation was only detected at 16 weeks of HFD and did not correlate with the induction of insulin resistance. CONCLUSIONS/INTERPRETATION: HFD-induced whole-body insulin resistance is initiated by impaired hepatic insulin action and exacerbated by skeletal muscle insulin resistance and is associated with the accumulation of specific bioactive lipid species.

Interleukin-6-deficient mice develop hepatic inflammation and systemic insulin resistance.

AIMS/HYPOTHESIS: The role of IL-6 in the development of obesity and hepatic insulin resistance is unclear and still the subject of controversy. We aimed to determine whether global deletion of Il6 in mice (Il6 (-/-)) results in standard chow-induced and high-fat diet (HFD)-induced obesity, hepatosteatosis, inflammation and insulin resistance. METHODS: Male, 8-week-old Il6 (-/-) and littermate control mice were fed a standard chow or HFD for 12 weeks and phenotyped accordingly. RESULTS: Il6 (-/-) mice displayed obesity, hepatosteatosis, liver inflammation and insulin resistance when compared with control mice on a standard chow diet. When fed a HFD, the Il6 (-/-) and control mice had marked, equivalent gains in body weight, fat mass and ectopic lipid deposition in the liver relative to chow-fed animals. Despite this normalisation, the greater liver inflammation, damage and insulin resistance observed in chow-fed Il6 (-/-) mice relative to control persisted when both were fed the HFD. Microarray analysis from livers of mice fed a HFD revealed that genes associated with oxidative phosphorylation, the electron transport chain and tricarboxylic acid cycle were uniformly decreased in Il6 (-/-) relative to control mice. This coincided with reduced maximal activity of the mitochondrial enzyme ß-hydroxyacyl-CoA-dehydrogenase and decreased levels of mitochondrial respiratory chain proteins. CONCLUSIONS/INTERPRETATION: Our data suggest that IL-6 deficiency exacerbates HFD-induced hepatic insulin resistance and inflammation, a process that appears to be related to defects in mitochondrial metabolism.

Mycobactericidal activity of selected disinfectants using a quantitative suspension test.

In this study, a quantitative suspension test carried out under both clean and dirty conditions was used to assess the activity of various instrument and environmental disinfectants against the type strain NCTC 946 and an endoscope washer disinfector isolate of Mycobacterium chelonae, Mycobacterium fortuitum NCTC 10,394, Mycobacterium tuberculosis H37 Rv NCTC 7416 and a clinical isolate of Mycobacterium avium-intracellulare (MAI). The disinfectants tested were; a chlorine releasing agent, sodium dichloroisocyanurate (NaDCC) at 1000 ppm and 10,000 ppm av Cl; chlorine dioxide at 1100 ppm av ClO2 (Tristel, MediChem International Limited); 70% industrial methylated spirits (IMS); 2% alkaline glutaraldehyde (Asep, Galan); 10% succinedialdehyde and formaldehyde mixture (Gigasept, Schulke & Mayr); 0.35% peracetic acid (NuCidex, Johnson & Johnson); and a peroxygen compound at 1% and 3% (Virkon, Antec International). Results showed that the clinical isolate of MAI was much more resistant than M. tuberculosis to all the disinfectants, while the type strains of M. chelonae and M. fortuitum were far more sensitive. The washer disinfector isolate of M. chelonae was extremely resistant to 2% alkaline activated glutaraldehyde and appeared to be slightly more resistant than the type strain to Nu-Cidex, Gigasept, Virkon and the lower concentration of NaDCC. This study has shown peracetic acid (Nu-Cidex), chlorine dioxide (Tristel), alcohol (IMS) and high concentrations of a chlorine releasing agent (NaDCC) are rapidly mycobactericidal. Glutaraldehyde, although effective, is a slow mycobactericide. Gigasept and Virkon are poor mycobactericidal agents and are not therefore recommended for instruments or spillage if mycobacteria are likely to be present.

Evaluation of the antimicrobial activity of a new super-oxidized water, Sterilox, for the disinfection of endoscopes.

The antimicrobial activity of a new super-oxidized water, Sterilox, has been tested against Mycobacterium tuberculosis, Mycobacterium avium-intracellulare, Mycobacterium chelonae, Escherichia coli (including type O157), Enterococcus faecalis, Pseudomonas aeruginosa, Bacillus subtilis var niger spores, methicillin-resistant Staphylococcus aureus, Candida albicans, poliovirus type 2 and human immunodeficiency virus HIV-1. Under clean conditions, freshly generated Sterilox was found to be highly active against all these micro-organisms giving a 5 log10 (99.999%) or greater reduction in two minutes or less.

An evaluation of the suitability of the European suspension test to reflect in vitro activity of antiseptics against clinically significant organisms.

The effectiveness of four antiseptics representing soluble phenolics (Dettol), Quaternary Ammonium Compounds (QAC) (Dettol Hospital Concentrate: DHC), mixed QAC/chlorhexidine (Hibicet Hospital Concentrate: HHC) and povidone iodine (Betadine) was assessed using the proposed phase 2 step 1 European Suspension test. The in vitro activity of the antiseptics against two of the proposed challenge strains, i.e. Staphylococcus aureus and Pseudomonas aeruginosa, was compared with that of 14 problematic clinical isolates of bacteria from a range of genera, including some multiple antibiotic resistant strains, and a clinical isolate of Candida albicans. In addition to the 5 min contact time recommended in the European test, a 1 min time was included. All four products, at their recommended use dilutions and a contact time of 5 min, achieved a Microbicidal Effect (ME) log reduction of at least 5 against the majority of organisms. Differences in activity between products were more pronounced and therefore the tests more discriminatory, when the contact time was reduced to 1 min. The clinical strains were not overtly more resistant to antiseptics than the standard test strains, suggesting that the CEN test strains mimic the antiseptic susceptibility of clinical isolates.

Mycobacterium terrae: a potential surrogate for Mycobacterium tuberculosis in a standard disinfectant test.

The susceptibility of Mycobacterium tuberculosis and Mycobacterium avium-intracellulare to the disinfections used for spillage and heat sensitive instruments has received much attention in recent years. The use of clinical isolates of M. tuberculosis and M. avium-intracellulare as test organisms is considered unsuitable for standard tests due to their hazardous nature (category 3 pathogens and slow growth rates). This has led to much debate in standards committees on the selection and use of a possible surrogate which would be safer and more practical to use and yet mimic the susceptibility of clinical isolates. This study compared the susceptibility of one possible surrogate Mycobacterium terrae NCTC 10856, with that of clinical isolates of M. tuberculosis H37 Rv and M. avium-intracellulare using a quantitative suspension test. The instrument and environmental disinfectants tested were a chlorine-releasing agent, sodium dichloroisocyanyurate (NaDCC) at 1000 ppm and 10,000 ppm av. Cl, chlorine dioxide at 1100 ppm av. ClO2 (Tristel, HayMan MediChem), 0.35% peracetic acid (NuCidex, Johnson & Johnson), 70% industrial methylated spirit (IMS), 2% alkaline glutaraldehyde (Asep, Galen), 10% succine dialdehyde and formaldehyde mixture (Gigasept, Schulke and Mayr). Results showed that the clinical isolate of M. avium-intracellulare was the most resistant of the three test organisms. M. terrae, which is not a category 3 pathogen, was slightly more resistant than M. tuberculosis and this would appear to be a suitable surrogate for establishing tuberculocidal activity. However, with an increase in the clinical significance of M. avium-intracellulare, particularly in human immunodeficiency virus (HIV) and immunocompromised patients, a more resistant surrogate is required. In the absence of such a surrogate, testing with M. avium-intracellulare in a clinical laboratory equipped for handling category 3 pathogens is still advised to establish mycobactericidal activity.

Glutaraldehyde-resistant Mycobacterium chelonae from endoscope washer disinfectors.

Glutaraldehyde is used to disinfect flexible and other heat-sensitive endoscopes often with the aid of automated systems. Mycobacterium chelonae is being isolated with increasing frequency from these washer disinfectors and processed endoscopes. This has, on occasions, led to misdiagnosis and iatrogenic infections. Recent reports suggest that disinfecting machines, on a sessional or regular basis, with 2% glutaraldehyde may have selected and therefore encouraged the growth of strains of Myco. chelonae, possibly in biofilm, with decreasing susceptibility to glutaraldehyde. In view of this, the resistance of three strains of Myco. chelonae var. chelonae (the type strain NCTC 946 and two machine isolates) was tested against 2% glutaraldehyde and a wide range of alternative disinfectants. Disinfectants tested were a chlorine releasing agent, sodium dichloroisocyanurate at 1000 ppm and 10,000 ppm av Cl, 0.35% peracetic acid (NuCidex, Johnson & Johnson), 70% industrial methylated spirit (IMS), 1% peroxygen compound ('Virkon', Antec International) and 10% succine dialdehyde ('Gigasept', Sanofi Winthrop). Suspension and carrier tests were carried out in the presence and absence of an organic load. Results showed the type strain, which had not been exposed to the selective pressure of disinfectant usage, to be very sensitive to most disinfectants with the exception of 1% Virkon. The washer disinfector isolates, on the other hand, were extremely resistant to 2% glutaraldehyde and showed greater resistance to 1% Virkon and 1000 ppm NaDCC. Purchasing machines in which the entire fluid pathways, including those for delivering rinse water, are disinfected with an appropriate agent during each cycle are preferred. If this is not possible then sessional cleaning and disinfection at the start of each day and regular maintenance should prevent biofilm formation and contamination with disinfectant-resistant strains of mycobacteria. In addition to machine disinfection, the use of sterile or bacteria-free (filtered < 0.45 microm) water is essential for bronchoscopes and all invasive endoscopes. If there is doubt that the effectiveness of the machine disinfection procedure or water quality, the channels and surfaces of endoscopes may be rinsed with 70% IMS after automated processing.

Risk of airborne transmission in an operating theatre containing four ultraclean air units.

This study shows that a single, large, operating theatre (barn) containing four ultraclean operating units (cabins), was highly effective in reducing the number of airborne bacteria in the operating fields providing all occupied ultraclean cabins were functioning correctly. The air flows and bacterial counts during operations within the cabins met the current standard for ultraclean systems (HTM 2025 1994) and there was no evidence of mixing of air between cabins. It is, however, recommended that air flows are regularly checked for compliance with the standard. If failure occurs in any single ultraclean unit, surgery in that cabin should cease as contaminated air may enter from the barn and surrounding cabins. Routine microbiological sampling should not be necessary providing there is no evidence of filter leakage. An operating theatre with several ultraclean operating tables in a single room would appear to be a viable proposition for the future. Considerable savings are likely in revenue costs as much of the air is reused and support services are shared.

Endoscope decontamination: automated vs. manual.

Automated endoscope washer disinfectors are widely used for the decontamination of flexible endoscopes. They are more effective than manual techniques and reduce the likelihood of skin contact with irritant disinfectants. Suitable machines are those which effectively clean, disinfect and rinse all channels and external surfaces without damaging the instrument. If glutaraldehyde is used, fumes should be removed or contained to protect endoscopy and processing staff. Machines should also be equipped with a self-disinfect facility and the rinse water should be of a suitable microbiological quality for the instruments processed, i.e. bacteria-free (sterile or filtered) water should be used for bronchoscopes and all invasive endoscopes. The choice of machine and cycle will depend on the following: whether a mobile or fixed unit is required; the type of disinfectant used; instrument throughput; and whether or not it is necessary to process more than one endoscope at a time. Purchasers are advised to request independent test reports which substantiate manufacturers' claims.

Endoscope decontamination: where do we go from here?

Thorough cleaning and disinfection or sterilization of endoscopes and associated equipment will reduce the likelihood of misdiagnosis and post-procedural infection. It will also prevent instrument deterioration and malfunction. With a rapid escalation in demand for endoscopy, particularly that associated with minimally invasive surgery, it is important that we have the processing technology to match the diagnostic and therapeutic value of these instruments without exposing staff and patients to unnecessary risk. Wherever possible staff should purchase heat tolerant endoscopic equipment that is readily accessible for cleaning. Automated processors, e.g. washer disinfectors and ultrasonic cleaners, improve the quality of the decontamination process but machines must have a self-disinfect function to prevent instrument recontamination during processing. Sterile, or filtered bacteria-free, water is essential for bronchoscopes and invasive instruments. Glutaraldehyde is still the most widely used disinfectant, particularly for the heat sensitive flexible endoscopes, but it is irritant and sensitizing and a safer alternative is sought. Peracetic acid is more rapidly efficacious and probably less irritant and, provided it does not damage endoscopes and processing equipment, may prove a suitable alternative. Unfortunately there are no nationally agreed test methods for assessing this and other new endoscope disinfectants and therefore no register of suitable or approved products. There is also no proven safe alternative to ethylene oxide for sterilizing invasive heat labile flexible endoscopes. It is important that, if toxic disinfectants and sterilants are used, staff and patients are suitably protected from exposure. Update training is essential for all processing staff if infection risks are to be minimized and sensitization problems avoided.

Infection risk and potential contamination of urine specimens associated with sample port design of catheter leg bags.

The ease of disinfection of the sample ports of three types of urine drainage leg bags with different sampling port systems was assessed using a bladder bag model. The ports were contaminated with Escherichia coli, 'disinfected' using a standard method, then sampled at time intervals up to one week after contamination. It was discovered that leg bags which employ a needle-based sampling system (the 'sample safe port system') were easier to disinfect than those which did not, and that organisms are retained in large enough numbers to lead to misdiagnosis of a urinary tract infection or to pose a retrograde infection risk.

Evaluation of the Steris System 1 Peracetic Acid Endoscope Processor.

An automated endoscope sterilizing machine, the Steris System 1 Processor, was tested for bactericidal and sporicidal efficacy. The disinfectant, peracetic acid, was diluted to 0.2% within an enclosed system. The exposure time to the disinfectant was 12 min and the overall cycle time ranged from 25-38 min, mean 29 min. Preliminary suspension tests, with and without yeast or serum, showed a log10 reduction of > 5 with Pseudomonas aeruginosa, Staphylococcus aureus and Bacillus subtilis in 5 min with 0.2% peracetic acid. After a routine cycle in the machine, endoscopes contaminated with the same organisms showed no growth. Two of 24 spore strips, containing 10(6) B. subtilis showed a small number of survivors (less than 10 per strip). No significant damage to the endoscope was observed although the number of cycles tested was small (i.e. 31). The advantage of the system is that staff are not directly exposed to the agent, but the costs per cycle are higher than glutaraldehyde, since peracetic acid is not renewed. Unlike other automated processors the Steris machine has no cleaning cycle.

Methods of cleaning and disinfection.

Most equipment associated infection is due to inadequate cleaning and disinfection and not a failure in sterilisation practices. The method of disinfection chosen should depend on the risks associated with the procedure undertaken, the heat, pressure and chemical tolerances of the item and the time available for processing. Heat disinfection or sterilisation is preferred but if the item is heat sensitive, chemicals may have to be used. It is important that the process selected is effective against patient associated organisms and opportunistic pathogens present in the environment. The most effective stage of any decontamination procedure is thorough cleaning and this should accompany or precede all disinfection procedures. Automated processors offer the safest, most reliable option, providing they are suitably monitored and maintained and that staff wear appropriate protective clothing.

Preliminary study of test methods to assess the virucidal activity of skin disinfectants using poliovirus and bacteriophages.

Two tests for assessing the virucidal activity of antiseptics are proposed. These involve applying either poliovirus (vaccine strain Sabin 1 an) or Escherichia coli bacteriophage (MS2 or K1-5) to the fingertips. Both test viruses are considered safe although poliovirus may be unacceptably tolerant to antiseptics. The use of bacteriophages as test organisms precludes the need for sophisticated recovery systems and can be undertaken readily by any bacteriology laboratory. The virucidal activity of 70%, 80% and 90% ethanol, 7.5% povidone-iodine, and soap and water was assessed using these tests. Thorough cleansing, followed by disinfection with 90% ethanol, was the most effective treatment. Removal of viruses from the gloved hand was also assessed and this was found to be more easily achieved than cleaning and disinfecting the ungloved hand. Wearing gloves protects the hands from viral contamination but changing them after each patient or contact is expensive.

A test for the assessment of 'hygienic' hand disinfection using rotavirus.

A standardized test procedure is described in which finger tips are inoculated with bovine rotavirus. The level of virus recovered after disinfection of artificially contaminated hands with various disinfectant detergents, alcoholic solutions and alcoholic formulations was determined. The method was found to be easy to perform and reproducible. The most efficient method for removal of virus from fingertips was found to be treatment with alcoholic solutions or products. Soap and water and disinfectant detergents were found to be a much less effective method of removing virus from contaminated hands.

A test procedure for evaluating surgical hand disinfection.

A technique for assessing the immediate and prolonged efficacy of surgical scrubs and alcoholic hand rubs is described. A mean baseline count is obtained from all volunteers and logarithmic reductions in resident skin flora immediately after one or more applications, and after wearing gloves for 3 h, are measured. Loose-fitting surgical gloves are used for sampling resident flora. Preparations were applied using a standard technique for 2 min, apart from one test with 70% isopropanol (IPA) in which the application time was 30 s. Two studies are described, one of which compared four chlorhexidine scrubs, and the second 70% IPA, 7.5% povidone-iodine scrub, 2% triclosan cleanser and unmedicated bar soap. In spite of their constituent similarity, the four chlorhexidine scrubs varied considerably in efficacy and user acceptability. A 2 min application of 70% IPA was the most effective treatment, and gave log10 reductions of 1.65 for immediate and 1.58 for prolonged effect. This was marginally more effective than a 30 s application, but the difference was not significant. 'Hibiscrub' was the most effective aqueous formulation and gave reductions of 1.01 for immediate effect and 1.16 for prolonged effect. The test described could be used by reference centres and manufacturers to assess the efficacy of new and existing surgical hand disinfection formulations.