Evaluation of PCR in the molecular diagnosis of endocarditis

Objective. Infective endocarditis (IE) is diagnosed by the Duke criteria, which can be inconclusive particularly when blood cultures are negative. This study investigated the application of polymerase chain reaction (PCR) to identify bacterial DNA in excised valvular tissue, and its role in establishing the diagnosis of IE. Methods. Ninety-eight patients undergoing valve replacement surgery were studied. Twenty-eight patients were confirmed as definite for endocarditis by the Duke criteria; nine were considered as possible and 61 had no known or previous microbial infection of the endocardium. A broad-range PCR technique was used to amplify prokaryotic 16S rRNA genes present within homogenised heart valve tissue. Subsequent DNA sequencing of the PCR amplicon allowed identification of the infecting microorganism. Results. PCR results demonstrated the presence of bacterial DNA in the heart valves obtained from 14 out of 20 (70%) definite IE patients with positive blood cultures preoperatively. The causative microorganism for one patient with definite culture negative endocarditis was identified by PCR. Two out of nine (22%) of the valves from possible endocarditis patients also had bacterial DNA present converting them into the definite criteria whereas in the valves of seven out of nine (78%) of these patients no bacterial DNA was detected. Conclusion. The application of PCR to the explanted valves in patients with possible or confirmed diagnosis can augment the Duke criteria thereby improving post-surgical antimicrobial therapeutic options. © 2003 The British Infection Society. Published by Elsevier Ltd. All rights reserved.

Association between sciatica and Propionibacterium acnes

We hypothesised that the inflammation seen around the nerve root in patients with sciatica may be caused by microbial infection. We used a newly developed serological test to diagnose deep-seated infections caused by low virulent gram-positive microorganisms. 43 of 140 (31%) patients with sciatica tested positive. Intervertebral disc material from a further 36 patients with severe sciatica who had undergone microdiscectomy was cultured for the presence of microorganisms. 19 of these patients (53%) had positive cultures after long-term incubation. Propionibacterium acnes was isolated from 16 of the 19 (84%) positive samples. Low virulent microorganisms, in particular P acnes, might be causing a chronic low-grade infection in the lower intervertebral discs of patients with severe sciatica. These microorganisms could have gained access to the spinal disc after previous minor trauma.

Infection risk associated with a closed luer access device

The potential for microbial contamination associated with a recently developed needleless closed luer access device (CLAD) (Q-Syte™ Becton Dickinson, Sandy, UT, USA) was evaluated in vitro. Compression seals of 50 multiply activated Q-Syte devices were inoculated with Staphylococcus epidermidis NCTC 9865 in 25% (v/v) human blood and then disinfected with 70% (v/v) isopropyl alcohol followed by flushing with 0.9% (w/v) sterile saline. Forty-eight of 50 (96%) saline flushes passed through devices that had been activated up to a maximum of 70 times remained sterile. A further 25 Q-Syte CLADs that had undergone multiple activations were challenged with prefilled 0.9% (w/v) sterile saline syringes, the external luer tips of which had been inoculated with S. epidermidis NCTC 9865 prior to accessing the devices. None of the devices that had been accessed up to 70 times allowed passage of micro-organisms, despite challenge micro-organisms being detected on both the syringe tip after activation and the compression seals before decontamination. These findings suggest that the Q-Syte CLAD may be activated up to 70 times with no increased risk of microbial contamination within the fluid pathway. The device may also offer protection from the external surface of syringe tips contaminated with micro-organisms. © 2005 Hospital Infection Society. Published by Elsevier Ltd. All rights reserved.

The microbial diagnosis of infective endocarditis

This review suggests an evidence-based algorithm for sequential testing in infective endocarditis. It discusses blood culture and the merits and drawbacks of serology in making the diagnosis. Newer techniques are briefly reviewed. The proposed algorithm will complement the Duke criteria in clinical practice. © 2003 The British Infection Society. Published by Elsevier Science Ltd. All rights reserved.

The source, cost and prevention of central venous catheter-related infection

Central venous catheters have become an integral part of patient management however they are associated with many complications including infection. Despite efforts being made to reduce the incidence of such infect ions the problem continues to increase and has resource implications for the Health Service. Studies relating to the source of microorganisms causing CVC-associated infection, the cost of such infections and the efficacy of an antimicrobial catheter have been undertaken. Thirty patients who required a CVC as part of their medical management and underwent cardiac surgery had the distal tips of their catheters sampled whilst in situ. Sampling took place within 1 h of catheter placement. Bacteria were isolated from 16% of the catheter distal tips sampled in situ. The guidewires used to insert the devices were also contaminated (50%). When CVC were inserted via a protective sheath, avoiding contact with the skin. the incidence of microbial contamination was reduced. These findings suggest that despite rigorous skin disinfection and strict aseptic technique, viable microorganisms are impacted onto the distal tip of CVC during the insertion procedure. Needleless intravascular access devices have been introduced in order to reduce the incidence of need1estick injury. However, it was unclear whether such connectors would act as a portal of entry for microorganisms to CVC. The efficacy of these devices was investigated. Within the controlled laboratory environment it was demonstrated that needleless devices, when challenged with microorganisms, did not allow the passage of microbes when flu id was injected. This therefore suggested that the devices should not increase the risk of catheter colonisation. When used in clinical practice however microbial contamination of the needleless connectors was 55 % in comparison to the routinely used luer connectors (23%). The cost of infections associated with CVC was determined. Twenty patients catheterised with a CVC designed for long term use who were admitted to hospital with a presumptive diagnosis of catheter-related infection were studied. The treatment given specifically for this infection was costed. The mean cost of such an infection was £ 1781.81. Throughout the UK this may amount to £1.565.906 per annum. The cost of infections associated with CVC designed for short term use was estimated to be between 5 and 7 million pounds per annum in the UK. In an attempt to reduce both the incidence and cost of catheter- related infection antimicrobial CVC have been developed. The efficacy of a novel polyurethane CVC impregnated on both the internal and external catheter surface with the quaternary ammonium compound benzalkonium chloride was investigated. Eighty eight patients received an antimicrobial catheter and 78 patients a conventional polyurethane CVC. The anti-microbial CVC resulted in a reduction in microbial colonisation of the external and internal polymer surfaces as compared to the control device. The observed reduction in microbial colonisation with the anti-microbial CVC may decrease the likelihood of subsequent infection offering a useful approach to the prevention of catheter-related infections.

Influence of microbial antigen formulation and delivery route on the immune response

Recent technological advances have resulted in the production of safe subunit and synthetic small peptide vaccines. Unfortunately, these vaccines are weakly or non-immunogenic in the absence of an immunological adjuvant (agents that can induce strong immunity to antigens). In addition, in order to prevent and/or control infection at the mucosal surface, stimulation of the mucosal immune system is essential. This may be achieved via the common mucosal immune system by exposure to antigen at a mucosal surface remote from the area of infection. Initial studies investigated the potential of multiple emulsions in effecting oral absorption and the subsequent immune responses to a lipopolysaccharide vaccine (LPS) after immunisation. Nasal delivery of LPS was carried out in parallel work using either aqueous solution or gel formulations. Tetanus toxoid vaccine in simple solution was delivered to guinea pigs as free antigen or entrapped in DSPC liposomes. In addition, adsorbed tetanus toxoid vaccine was delivered nasally free or in an aerosil gel formulation. This work was extended to investigate guinea pigs immunised by various mucosal routes with a herpes simplex virus subunit vaccine prepared from virus infected cells and delivered in gels, multiple emulsions and liposomes. Comparable serum antibody responses resulted but failed to produce enhanced protection against vaginal challenge when compared to subcutaneous immunisation with alhydrogel adjuvanted vaccine. Thus, immunisation of the mucosal surface by these methods may have been inadequate. These studies were extended in an attempt to protect against HSV genital challenge by construction of an attenuated Salmonella typhimurium HWSH aroA mutant expressing a cloned glycoprotein D-l gene fused to the Es-cherichia coli lac z promoter. Preliminary work on the colonisation of guinea pigs with S. typhimurium HWSH aroA mutants were carried out, with the aim of using the guinea pig HSV vaginal model to investigate protection.

A randomized, prospective clinical trial to assess the potential infection risk associated with the PosiFlow® needleless connector

The microbial contamination rate of luers of central venous catheters (CVCs) with either PosiFlow® needleless connectors or standard caps attached was investigated. The efficacy of 70% (v/v) isopropyl alcohol, 0.5% (w/v) chlorhexidine in gluconate 70% (v/v) isopropyl alcohol and 10% (w/v) aqueous povidone-iodine to disinfect the intravenous connections was also assessed. Seventy-seven patients undergoing cardiac surgery who required a CVC as part of their clinical management were randomly allocated either needleless connectors or standard caps. Patients were also designated to receive chlorhexidine/alcohol, isopropyl alcohol or povidone-iodine for pre-CVC insertion skin preparation and disinfection of the connections. After 72 h in situ the microbial contamination rate of 580 luers, 306 with standard caps and 274 with needleless connectors attached, was determined. The microbial contamination rate of the external compression seals of 274 needleless connectors was also assessed to compare the efficacy of the three disinfectants. The internal surfaces of 55 out of 306 (18%) luers with standard caps were contaminated with micro-organisms, whilst only 18 out of 274 (6.6%) luers with needleless connectors were contaminated (P<0.0001). Of those needleless connectors disinfected with isopropyl alcohol, 69.2% were externally contaminated with micro-organisms compared with 30.8% disinfected with chlorhexidine/alcohol (P<0.0001) and 41.6% with povidone-iodine (P<0.0001). These results suggest that the use of needleless connectors may reduce the microbial contamination rate of CVC luers compared with the standard cap. Furthermore, disinfection of needleless connectors with either chlorhexidine/alcohol or povidone-iodine significantly reduced external microbial contamination. Both these strategies may reduce the risk of catheter-related infections acquired via the intraluminal route. © 2003 The Hospital Infection Society. Published by Elsevier Science Ltd. All rights reserved.

A prospective clinical trial to evaluate the microbial barrier of a needleless connector

Needleless connectors are being increasingly used for direct access to intravascular catheters. However, the potential for microbial contamination of these devices and subsequent infection risk is still widely debated. In this study the microbial contamination rate associated with three-way stopcock luers with standard caps attached was compared to those with Y-type extension set luers with Clearlink® needleless connectors attached. Fifty patients undergoing cardiothoracic surgery who required a central venous catheter (CVC) as part of their peri- and postoperative management were studied for microbial contamination of CVC luers following 72 hrs in situ. Each patient's CVC was randomly designated to have either the three-way stopcocks with caps (control patients) or Clearlink® Y-type extension sets (test patients). Prior to, and following each manipulation of the three-way stopcock luers or Clearlink® devices, a 70% (v/v) isopropyl alcohol swab was used for disinfection of the connections. The microbial contamination of 393 luers, 200 with standard caps and 193 with Clearlink® attached, was determined. The internal surfaces of 20 of 200 (10%) three-way stopcock luers with standard caps were contaminated with micro-organisms whereas only one of 193 (0.5%) luers with Clearlink® attached was contaminated (P < 0.0001). These results demonstrate that the use of the Clearlink® device with a dedicated disinfection regimen reduces the internal microbial contamination rate of CVC luers compared with standard caps. The use of such needle-free devices may therefore reduce the intraluminal risk of catheter-related bloodstream infection and thereby supplement current preventive guidelines. © 2006 The Hospital Infection Society.

A comparative study to evaluate surface microbial contamination associated with copper-containing and stainless steel pens used by nurses in the critical care unit

A clinical study was undertaken to compare the surface microbial contamination associated with pens constructed of either a copper alloy or stainless steel used by nurses on intensive care units. A significantly lower level of microbial contamination was found on the copper alloy pens. Copyright © 2011 by the Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.