Role of bacterial adherence and the mucus barrier on bacterial translocation: effects of protein malnutrition and endotoxin in rats.

OBJECTIVE: The purpose of the study was to investigate the potential relations between mucosal bacterial adherence, intestinal mucus and mucin content, and bacterial translocation. SUMMARY BACKGROUND DATA: The attachment of bacteria to mucosal surfaces is the initial event in the pathogenesis of most bacterial infections that originate at mucosal surfaces, such as the gut. The intestinal mucus layer appears to function as a defensive barrier limiting micro-organisms present in the intestinal lumen from colonizing enterocytes. Consequently, studies focusing on the biology of bacterial adherence to the intestinal mucosa likely are to be important in clarifying the pathogenesis of gut origin sepsis. METHODS: To explore the relations between intestinal bacterial adherence, mucus bacterial binding, and bacterial translocation, two models were used. One (protein malnutrition) in which profound alterations in intestinal morphology occurs in the absence of significant translocation and one (endotoxin challenge) in which bacterial translocation occurs and intestinal morphology is relatively normal. RESULTS: Protein malnutrition was not associated with bacterial translocation and measurement of enteroadherent, mucosally associated bacterial population levels documented that the total number of gram-negative enteric bacilli adherent to the ileum and cecum was less in the protein-malnourished rats than in the normally nourished animals (p < 0.01). Furthermore, there was an inverse relation between the duration of protein malnutrition and bacterial adherence to the intestinal mucosa (r = 0.62, p < 0.002). In contrast, after endotoxin challenge, the level of enteroadherent bacteria was increased and bacterial translocation was observed. The binding of Escherichia coli to immobilized ileal mucus in vitro was decreased significantly in protein-malnourished rats, whereas E. coli binding to insoluble ileal mucus was increased in the rats receiving endotoxin. CONCLUSIONS: This study indicates that the adherence of bacteria to the intestinal mucosal surface is an important factor in bacterial translocation, that intestinal mucus modulates bacterial adherence, and that increased levels of mucosally associated bacteria are associated with a loss intestinal barrier function to bacteria.     

Successful Lung Re-transplant in a Patient with Cepacia Syndrome due to Burkholderia ambifaria

Chronic airway inflammation and infection drive morbidity and mortality among patients with cystic fibrosis (CF). While Haemophilus influenzae and Staphylococcus aureus predominate in children, the prevalence of Pseudomonas aeruginosa increases as patients age. Other bacteria, including species within the Burkholderia cepacia complex (Bcc), are also more prevalent among adults with CF. Species within the Bcc accelerate lung function decline and can trigger development of “cepacia syndrome,” both before and after lung transplantation. As a result, some centers advise against lung transplantation for Bcc-infected patients; however, little is known about the relative virulence of uncommon Bcc species. We describe a successful lung re-transplant in a patient with CF, chronic Burkholderia ambifaria airway infection, and cepacia syndrome.     

Reduction of bacterial adherence to catheter surface with heparin

Despite many advances in catheter design and use, the most common cause of hospital-acquired infections is catheterization of the urinary tract. In the present investigation the adherence of bacteria to catheters coated with heparin was studied. Since heparin itself does not coat the plastic catheter surface, a complex of heparin with tridodecylmethylammonium chloride (TDMAC) was used which results in hydrophobic association of hydrocarbon chains of the TDMAC with the catheter leaving the quaternary ammonium moiety of TDMAC exposed to the surface forming ionic bonds with the highly anionic sulfate groups of heparin. Coating latex catheter material with TDMAC without heparin resulted in 3.6-fold higher adherence whereas coating with the TDMAC-heparin complex reduced adherence to less than 10% of control untreated latex. TDMAC-heparin also significantly reduced bacterial adherence to teflon coated latex (Bardex) and vinyl catheter material. Less than 30% of the original heparin was removed after wash periods of up to one week. These results indicate that TDMAC-heparin coating of urethral catheters reduces bacterial adherence and thereby may delay the acquisition of catheter associated urinary tract infection.     

Halothane decreases bacterial adherence in vitro

BACKGROUND: Adherence of pathogenic bacteria to host epithelial cells is thought to be the initial step in infection, while the presence of the commensal flora is an important host defence mechanism. Anything altering bacterial adherence to human epithelial cells may contribute to bacterial infections. The impact of anaesthesia on this first step to infection is not known. In this study the effect of halothane on bacterial adherence was investigated. METHODS: Human epithelial cells (HEp-2) and two strains of Escherichia coli were exposed to halothane 2% for 2 h. Then HEp-2 cells were coincubated with bacteria for 3 h. Bacteria attached to the epithelial cells were evaluated by light microscopy. RESULTS: Compared to the control, bacterial adherence was reduced by 37% to 56% with the different strains when HEp-2 cells were exposed to halothane. No significant difference was found when only bacteria were treated with halothane. CONCLUSION: Our results show that halothane reduces bacterial adherence to human epithelial cells in vitro. Reduced number or function of epithelial cell surface receptors may be responsible for the reduced adherence as no changes were observed when only the bacteria were exposed to halothane.     

Further characterization of bacterial adherence to urinary bladder mucosa: comparison with adherence to anion exchange resin

Bacterial adherence to anion exchange resin has recently been reported to provide a useful, rapid, in vitro screening assay for identifying putative antiadherence agents. The studies presented in this report provide additional evidence that adherence to anion exchange resin is similar to urinary bladder mucosa adherence. Results are as follows. 1) Heparin inhibits the adherence of Escherichia coli, Klebsiella ozaenae, Proteus mirabilis, Streptococcus fecalis and Pseudomonas aeruginosa to both the mucin deficient bladder and anion exchange resin. 2) Drugs which inhibit E. coli adherence to the bladder also inhibit E. coli adherence to the resin and conversely, drugs which do not effect E. coli adherence to the resin do not affect attachment to the bladder. 3) Extracts from mammalian urinary bladder mucosa prevent E. coli adherence to both the bladder and resin. 4) The ability of heparin to displace preattached bacteria from anion exchange resin is inversely proportional to the time the bacteria are in contact with resin. Similar results were obtained for the ability of heparin to displace bacteria from the bladder. These studies indicate that bacterial adherence to anion exchange resin responds to heparin and other chemical agents in a manner similar to the mucin deficient rabbit urinary bladder. Because of the ease and rapid nature of this in vitro assay, it serves as a useful screen for potential bacterial antiadherence agents and could be used to help elucidate mechanisms of bacterial attachment.     

Overrepresentation of immunostimulatory CpG motifs in Burkholderia genomes.

Pulmonary infections with Burkholderia cepacia complex organisms contribute significantly to morbidity and mortality in patients with cystic fibrosis (CF), partially due to the intense inflammatory response of the host to the presence of bacteria and their byproducts. In the present study we show that Burkholderia genomes contain a large number of immunostimulatory CpG motifs. This is mainly because of their large genome size. This suggests that DNA from Burkholderia sp. has the potential to cause significant inflammatory response. Whether this contributes significantly to the airway inflammation often observed in infected CF patients remains to be determined.     

Bacterial adherence to surgical sutures. A possible factor in suture induced infection.

Surgical sutures are known to potentiate the development of wound infection. The purpose of this study was to investigate whether the capability of bacteria to adhere to various types of sutures has a significant effect on their ability to cause infections. Bacterial adherence to sutures was quantitatively measured using radiolabeled bacteria. In vitro adherence assays revealed remarkable variations in the affinity of bacteria to the various sutures: nylon bound the least bacteria while bacterial adherence to braided sutures (silk, Ti-cron, Dexon) was five to eight folds higher. The degree of infection obtained in mice in the presence of different sutures nicely correlated with their adherence properties. The different removal rate of adherent bacteria (glutaraldehyde-fixed) from various sutures by the tissue factors in mice supports the hypothesis that bacterial adherence to suture materials plays a significant role in the induction of surgical infection. Our observation points out at the need for careful suture selection in contaminated wounds. The adherence properties of sutures should be considered in any future surgical suture design.     

In vitro adherence of bacteria to sutures in cardiac surgery

The adherence of bacteria to sutures used in cardiac surgery was studied by in vitro quantitative determination with [3H]-leucine-labeled Staphylococcus epidermidis, Staphylococcus aureus and Streptococcus sanguis. The adherence per unit area for staphylococci was least for monofilament polypropylene (Prolene), 3 times higher (p less than 0.05) for braided polyester (Mersiline) and greatest (10 times, p less than 0.005) for braided polyester sutures coated with polybutylate (Ethibond). Mean values for the adherence of streptococci were low for all the sutures. Sutures pretreated with human plasma showed a 12-37% increase in bacterial adherence. The cell surface hydrophobicity, surface charge and the haemagglutinating property of bacteria did not correlate with their adherence property. In view of these observations, it is suggested that: (a) the preferential adherence of staphylococci to intra-cardiac sutures may be one of the explanations for its being the commonest cause of early prosthetic valve endocarditis, (b) there is a need for a careful selection of sutures used in cardiac surgery and (c) the described in vitro assay for bacterial adherence may be used for monitoring the development of better designed sutures and the effect of incorporation of antibiotics in the sutures.     

Bacterial adherence to tantalum versus commonly used orthopedic metallic implant materials

OBJECTIVES: Evaluation of bacterial adhesion to pure tantalum and tantalum-coated stainless steel versus commercially pure titanium, titanium alloy (Ti-6Al-4V), and grit-blasted and polished stainless steel. DESIGN: Experimental in vitro cell culture study using Staphylococcus aureus and Staphylococcus epidermidis to evaluate qualitatively and quantitatively bacterial adherence to metallic implants. METHODS: A bacterial adhesion assay was performed by culturing S. aureus (ATCC 6538) and S. epidermidis (clinical isolate) for one hour with tantalum, tantalum-coated stainless steel, titanium, titanium alloy, grit-blasted and polished stainless steel metallic implant discs. Adhered living and dead bacteria were stained using a 2-color fluorescence assay. Adherence was then quantitatively evaluated by fluorescence microscopy and digital image processing. Qualitative adherence of the bacteria was analyzed with a scanning electron microscope. The quantitative data were related to the implant surface roughness (Pa-value) as measured by confocal laser scanning microscopy. RESULTS: Bacterial adherence of S. aureus varied significantly (p = 0.0035) with the type of metallic implant. Pure tantalum presented with significantly (p < 0.05) lower S. aureus adhesion compared to titanium alloy, polished stainless steel, and tantalum-coated stainless steel. Furthermore, pure tantalum had a lower, though not significantly, adhesion than commercially pure titanium and grit-blasted stainless steel. Additionally, there was a significantly higher S. aureus adherence to titanium alloy than to commercially pure titanium (p = 0.014). S. epidermidis adherence was not significantly different among the tested materials. There was no statistically significant correlation between bacterial adherence and surface roughness of the tested implants. CONCLUSIONS: Pure tantalum presents with a lower or similar S. aureus and S. epidermidis adhesion when compared with commonly used materials in orthopedic implants. CLINICAL IMPLICATION: Because bacterial adhesion is an important predisposing factor in the development of clinical implant infection, tantalum may offer benefits as an adjunct or alternative material compared with current materials commonly used for orthopedic implants.     

Successful treatment of cepacia syndrome with a combination of intravenous cyclosporin, antibiotics and oral corticosteroids

Burkholderia cepacia complex (BCC) is a group of 17 closely related bacterial species that can cause pulmonary infection in patients with cystic fibrosis (CF). The clinical manifestations of BCC infection are varied but can include cepacia syndrome, which is a rapidly progressing necrotising pneumonia with an almost universally fatal outcome. We report the case of a 38year old man, known to have chronic infection with the ET12 strain of Burkholderia cenocepacia who developed cepacia syndrome 26years after initial infection. Aggressive treatment with a combination of 4 intravenous antibiotics, oral corticosteroids and cyclosporin brought about clinical, radiological and biochemical resolution of his cepacia syndrome. This case highlights the possible role of cyclosporin in the treatment of cepacia syndrome.     

Inhibition of sodium urate crystal adherence to bladder surface by polysaccharide

Bladder surface glycosaminoglycan (polysaccharide) has been shown to act in a nonspecific manner as an antiadherence factor and may potentially be important in the pathogenesis (and prevention) of renal calculi. It has been shown that when present, bladder surface GAG will reduce the adherence of both calcium and calcium oxalate crystals. This study was conducted to see if a similar effect would be found with uric acid crystals, in that the presence of the normal bladder mucus with its contained GAG and exogenously supplied polysaccharide, pentosanpolysulfate, would be inhibitory to crystal adherence. Radioactively labeled uric acid crystals were made by dissolving 14C labeled sodium urate to a point of saturation at pH 7.5 and then rapidly lowering the pH to 5.5. These crystals were placed into four groups of bladders: 1) normal bladders, 2) bladders rendered mucus deficient with hydrochloric acid, 3) mucus deficient bladders which were pre-treated with sodium pentosanpolysulfate prior to crystal addition and 4) mucus deficient bladders to which were added crystals pretreated with pentosanpolysulfate. There was a significant rise in crystal adherence after acid treatment of the bladder, a 2.3-fold increase, which was blocked with prior treatment of the crystals with pentosanpolysulfate and to a lesser degree by pretreating mucus deficient bladders with pentosanpolysulfate.     

In vitro quantitative adherence of bacteria to intravascular catheters

Adapting standard techniques, a simple in vitro system was devised to compare quantitative bacterial adherence to iv catheters of different compositions. Upon brief immersion of catheters in suspensions of Staphylococcus aureus, coagulase-negative staphylococci, and Escherichia coli, organisms adhered to catheter surfaces. After overnight growth in broth, organisms remained adherent and formed colonies, as shown by light and scanning electron microscopy. In addition, quantitative adherence using a blood agar roll technique, expressed as bacteria per square centimeter of catheter surface area per 10(6) colonies per milliliter inoculum, was calculated. Adherence was greater on polyvinylchloride (PVC) catheters (geometric mean 342) than on Teflon catheters (geometric mean 49.6) for coagulase-negative staphylococci (P less than 0.005). Also, the number of coagulase-negative staphylococci adherent to PVC catheters was significantly greater than for E. coli (geometric mean 70.6) at analogous inocula (P less than 0.02). Differences in bacterial adherence to the surface of iv devices may be important in the pathogenesis of catheter-associated infections. This in vitro method could prove useful in testing bacterial adherence properties of newly developed catheter materials, and allow development of catheters less prone to be associated with bacterial adherence and catheter-related infections.     

Inhibition by immunoglobulins of Staphylococcus aureus adherence to fibronectin-coated foreign surfaces

Recent data suggest that fibronectin may favor Staphylococcus aureus infection by promoting attachment to either injured tissues or implanted foreign bodies. Using a previously described in vitro assay, we show that promotion of S. aureus adherence by surface-bound fibronectin, adsorbed on polymethylmethacrylate (PMMA) coverslips, is antagonized by antistaphylococcal antibodies present in immunoglobulin G (IgG) purified from human plasma. Among the different organisms tested, the protein A-deficient strain Wood 46 of S. aureus was the most strongly inhibited by purified IgG or whole serum dose-dependently. Bacterial adherence was not influenced by preincubating fibronectin-coated PMMA with either purified IgG or whole serum. However, inhibition of bacterial adherence was directly related to the extent of IgG binding to S. aureus Wood 46. When F(ab')2 fragments of purified IgG were tested in the adherence assay, they could also reduce the interaction between S. aureus Wood 46 and fibronectin-coated PMMA. Two other staphylococcal strains were also tested in the adherence inhibition assay: Whereas the protein A-rich strain Cowan I of S. aureus was moderately inhibited by purified IgG or whole serum, S. epidermidis KH 11 was not at all inhibited by IgG which bound poorly to the bacterial cells. This study has demonstrated that bacterial coating by humoral factors, and specifically IgG, may influence significantly subsequent adherence of S. aureus to surface-bound fibronectin.     

Bacterial adherence to vascular grafts after in vitro bacteremia

All currently used arterial prosthetics have a greater susceptibility to infection following bacteremia than does autogenous tissue. This experiment compares quantitative bacterial adherence to various prosthetic materials after bacteremia carried out in a tightly controlled and quantitative fashion. Ten centimeters long, 4 mm i.d. Dacron, umbilical vein (HUV), and polytetrafluoroethylene (PTFE) grafts, as well as PTFE grafts with a running suture line at the midportion were tested. Each graft was interposed into a pulsatile perfusion system modified from a Waters MOX 100 TM renal transplant pump. Indium-111-labeled Staphylococcus aureus were added to heparinized canine blood to give a mean concentration of 4.7 X 10(6) bacteria/cc. This infected blood was recirculated through each graft for 30 min at a rate of 125 cc/m, 100 Torr (sys), 60 beats/min. The gamma counts/graft were used to calculate the number of bacteria/cm2 of graft surface. After nine experiments, a mean of 9.63 X 10(5) bacteria/cm2 were adherent to the Dacron, 1.04 X 10(5) bacteria/cm2 to the HUV, and 2.15 X 10(4) bacteria/cm2 to the PTFE. These differences were all significant at the 0.05 level. The addition of a suture line increased bacterial adherence to the PTFE graft by 50%. These results suggest that PTFE is the vascular graft material of choice when a prosthetic graft must be implanted despite a high risk of subsequent clinical bacteremia. Our in vitro, pulsatile perfusion model gave accurate and reproducible results, and appears well suited for further studies of bacterial, or platelet adherence to grafts, as well as the biomechanics of vascular conduits.     

Staphylococcal adherence to chicken cartilage

Immature chicken cartilage was incubated in a Staphylococcus aureus suspension and then washed. Scanning electron microscopy and radiolabel measurements showed increased adherence to cartilage with increasing bacterial concentration. Preheating of the bacteria did not reduce the adherence property, but trypsin treatment did.     

Invasion and biofilm formation of Burkholderia dolosa is comparable with Burkholderia cenocepacia and Burkholderia multivorans.

BACKGROUND: Colonisation with Burkholderia cepacia complex pathogens has been associated with accelerated decline in cystic fibrosis (CF) patients. The two most common species among the CF community are Burkholderia cenocepacia and Burkholderia multivorans. However, Burkholderia dolosa has recently been causing concern due to its transmissibility and virulence in CF patients. METHODS: We have compared the ability of five B. dolosa strains to invade lung epithelial cells in vitro with other members of the Bcc. The bacterial epithelial cell interaction was visualised by transmission electron microscopy. We have also examined the ability of these strains to form biofilms in vitro. RESULTS: We have found that members of this species can invade pulmonary epithelial cells in vitro as readily as those from B. cenocepacia and B. multivorans. Confirmation of intracellular invasion was obtained by transmission electron microscopy. B. dolosa strains were readily observed in membrane bound vesicles inside the lung epithelial cells. In addition, strains from this species were capable of forming strong biofilms at a level comparable to the more clinically relevant species. CONCLUSIONS: B. dolosa shows comparable virulence characteristics in vitro to the two most clinically relevant species indicating precautions should be taken when this species is identified in the CF population.     

Staphylococcal adherence to chicken cartilage

Immature chicken cartilage was incubated in a Staphylococcus aureus suspension and then washed. Scanning electron microscopy and radiolabel measurements showed increased adherence to cartilage with increasing bacterial concentration. Preheating of the bacteria did not reduce the adherence property, but trypsin treatment did.     

Staphylococcal adherence to chicken cartilage

Immature chicken cartilage was incubated in a Staphylococcus aureus suspension and then washed. Scanning electron microscopy and radiolabel measurements showed increased adherence to cartilage with increasing bacterial concentration. Preheating of the bacteria did not reduce the adherence property, but trypsin treatment did.     

Bacterial adherence to vascular prostheses: A determinant of graft infectivity

An in vitro model was developed to quantitatively measure bacterial adherence to the surface of prosthetic vascular graft material. Four strains of bacteria (Staphylococcus aureus, nonmucin-producing S. epidermidis [SP-2], mucin-producing S. epidermidis [RP-12], and Escherichia coli) were used to inoculate expanded polytetrafluoroethylene (ePTFE), woven Dacron, and velour knitted Dacron graft material. After graft specimens were incubated in a 10⁷ suspension of bacteria, they were washed to remove nonadherent organisms and ultrasonically oscillated to dislodge adherent organisms. Quantitative culture of the sonication effluent was used to calculate bacterial adherence, expressed as the number of colony-forming units found in each square centimeter of graft material per 10⁷ inoculum. All bacterial strains had a greater affinity to velour knitted Dacron graft than to ePTFE (p < 0.025). E. coli and S. aureus adhered to velour knitted Dacron in greater numbers than to woven Dacron (p < 0.04). The production of extracellular polysaccharide (mucin) by the RP-12 strain significantly increased adherence to both ePTFE and Dacron grafts compared with the other three bacterial strains tested (p < 0.04). Although E. coli was less adherent to ePTFE than nonmucin-producing staphylococcal strains (S. aureus and SP-2), no difference in adherence to knitted or woven Dacron graft material was demonstrated. The differential adherence of bacteria to prosthetic vascular grafts pays an important role in the pathogenesis of graft sepsis and determines relative graft infectivity. The in vitro model developed is well suited for further study of the mechanisms by which bacteria adhere to and colonize vascular grafts. (J VASC SURG 1986;3:732-40.)     

Bacterial adherence to ofloxacin-blended polylactone-coated self-reinforced L-lactic acid polymer urological stents

OBJECTIVE: To determine whether ofloxacin coating has any effect on bacterial adherence to bioresorbable self-reinforced L-lactic acid polymer (SR-PLLA) urological stents. MATERIALS ANS METHODS: SR-PLLA stents were coated with epsilon-caprolactone/L-lactide copolymer blended with ofloxacin at three different concentrations of ofloxacin (0.5, 2 and 5% w/w). The adherence of five bacterial strains (Pseudomonas aeruginosa, Enterococcus faecalis, Proteus mirabilis and two strains of Escherichia coli) to the coated SR-PLLA stents was analysed. Uncoated stent pieces were used as controls. The effect of ofloxacin coating on bacterial growth in the microenvironment of the stent pieces was also analysed. RESULTS: Ofloxacin coating prevented bacterial adherence to SR-PLLA stent material; this effect correlated significantly with the ofloxacin concentration of the caprolactone coating. Ofloxacin coating reduced the amount of bacteria in the microenvironment of the stent, but because of natural resistance, ofloxacin coating had little effect on E. faecalis. CONCLUSION: Except for E. faecalis, ofloxacin coating may reduce stent-associated infections. However, further studies are needed to confirm its biocompatibility and efficacy in clinical use.