Investigation of the Actual Causes of Hip Joint Implant Loosening Classified as Aseptic--Analysis of Microbiological Culture Results and Levels of Inflammatory Markers.

Loosening of the hip joint prosthesis is considered as one of the most significant postoperative complications in recent years. The laboratory diagnostic procedure used to differentiate periprosthetic infection from aseptic loosening is very difficult because of the biofilm which microorganisms form on the implant surface. The purpose of this research was to evaluate the level of concordance between clinical classification of implant loosening among 50 patients subjected to reimplantation procedure and laboratory investigation of PJI including microbiological culture results and the levels of inflammatory markers assessed in the patients' synovial fluid samples, serum, and full blood. The synovial fluid was collected for leukocyte count, differential cell count, and culture on standard media. The levels of systemic inflammation markers such as the ESR and CRP concentration were determined in serum and full blood. Tissue samples were collected for microbiological studies. Components from endoprostheses were exposed to ultrasound in a process called sonication. Among the parameters measured in serum and full blood the levels of ESR and CRP were higher in the septic group of patients. Cytologic analysis of synovial fluid was in correlation with microbiologic identification. The most frequent isolated bacteria was Staphylococcus epidermidis. Culture results from materials such as synovial fluid, sonicate and tissues are crucial to establish the infectious aetiology of the loosening. Microscopic analysis of synovial fluid represents a simple, rapid and accurate method for differentiating PJI from aseptic failure. Sonication increases detection of the infectious process, and culture results are in correlation with the cytologic analysis of synovial fluid.

Sonication as a diagnostic approach used to investigate the infectious etiology of prosthetic hip joint loosening.

The purpose of the study was to evaluate the usefulness of sonication for the diagnosis of prosthetic joint infections (PJIs) by its comparison with periprosthetic tissues (PTs) and synovial fluid (SV-F) cultures. The study groups included 54 patients undergoing exchange of total hip prostheses for so called "aseptic" loosening occurring without clinical manifestations of an accompanying PJI and 22 patients who developed a sinus tract communicating with the prosthesis which was indicative of an ongoing infectious process. Significant positive culture results were obtained among 10 (18.5%) patients with "aseptic" implant failure and in 18 (81.8%) patients who developed a sinus tract. Sonicate-fluid (S-F) yielded bacterial growth in all culture-positive patients with "aseptic" loosening vs. 15 patients with presumed PJIs. There was a concordance in terms of bacterial species isolated from S-F and conventional cultures from individual patients. Coagulase-negative staphylococci were isolated most frequently. Sensitivity of sonication (75%) exceeded that estimated for PTs (69%) and SV-F (45%) cultures. We conclude that identification of causative agents of PJIs which is critical to further therapeutic decisions is aided by the combination of sonication and conventional culture.

Infectious prosthetic hip joint loosening: bacterial species involved in its aetiology and their antibiotic resistance profiles against antibiotics recommended for the therapy of implant-associated infections.

Reliable microbiological diagnosis along with surgery and prolonged antibiotic therapy are key elements in the management of prosthetic-joint infections (PJIs). The purpose of this study was to characterize antibiotic resistance profiles of bacteria involved in the aetiology of PJIs. A total of 33 bacterial isolates cultured from 31 patients undergoing exchange of total hip prostheses were analyzed. The diagnostic approach toward isolation of prosthesis- associated microorganisms included sonication of retrieved implants and conventional cultures of periprosthetic tissues and synovial fluid. The in vitro resistance profiles of bacterial isolates were determined in relation to antibiotics recommended for the therapy of PJIs using the disc diffusion method, E-tests(®) and broth microdilution system. Coagulase-negative staphylococci (CNS) were predominant microorganisms followed by Staphylococcus aureus, Enterobacter cloacae, Streptococcus mitis, and Propionibacterium acnes. Twenty out of 30 and 12 out of 30 staphylococcal isolates were methicillin- and multi-drug resistant, respectively. Only two isolates were rifampicinresistant. All staphylococci were susceptible to glycopeptides and linezolid. This paper stresses the pathogenic role of staphylococci in patients suffering from implant loosening and reports high methicillin- and multidrug-resistance rates in these bacteria. Hence, antimicrobial susceptibility tests of individual bacterial isolates must always be performed to guide selection of the optimal therapeutic option.

Characterization of Staphylococcus epidermidis and Staphyloccocus warneri small-colony variants associated with prosthetic-joint infections.

We determined the frequency of isolation of staphylococcal small-colony variants (SCVs) from 31 culture-positive patients undergoing revision of total hip prosthesis for aseptic loosening or presumed prosthetic-joint infection (PJI). We analysed auxotrophy of cultured SCVs, their antimicrobial susceptibility profiles and their biofilm-forming capacity. Eight SCV strains were cultivated from six (19 %) patients. All SCVs were coagulase-negative staphylococci (CNS) with Staphylococcus epidermidis as the predominant species; there was also one Staphylococcus warneri SCV. The SCVs were auxotrophic for haemin, with one strain additionally auxotrophic for menadione. We noted the presence of two phenotypically (differences concerning antimicrobial susceptibility) and genetically distinct SCV strains in one patient, as well as the growth of two genetically related SCVs that differed in terms of their morphology and the type of auxotrophy in another. Seven out of eight SCVs were resistant to meticillin and gentamicin. In addition, antibiotic sensitivity testing revealed three multidrug-resistant SCV-normal-morphology isolate pairs. One S. epidermidis SCV harboured icaADBC genes and was found to be a proficient biofilm producer. This paper highlights the involvement of CNS SCVs in the aetiology of PJIs, including what is believed to be the first report of a S. warneri SCV. These subpopulations must be actively sought in the routine diagnosis of implant-associated infections. Moreover, in view of the phenotypic and genetic diversity of some SCV pairs, particular attention should be paid to the investigation of all types of observed colony morphologies, and isolates should be subjected to antimicrobial susceptibility testing.

[Review of the effectiveness of an empirical antibiotic therapy in suspected ventilator-associated pneumonia]. / Ocena skutecznosci rekomendowanych scheatów antybiotykoterapii emprirycznej VAP w oparciu o analize in vitro lekowraziliwosci glównych patogenów.

BACKGROUND: Ventilator-associated pneumonia (VAP) occurs in approximately 10-20% of mechanically ventilated patients, and is associated with an extremely high mortality rate (up to 70%). The purpose of the study was to determine the susceptibility spectrum of Klebsiella, Pseudomonas and Acinetobacter strains isolated from VAP patients. METHODS: We analysed 81 strains of microorganisms isolated from bronchoalveolar lavages (BAL) of VAP patients. The minimal inhibitory concentrations (MIC) of antibiotics recommended for empirical therapy were determined using an automated VITEK 2 system, and for the MIC of doripenem - the Etest assay. Results were analysed following the guidelines of the Clinical and Laboratory Standards Institute. RESULTS: For infections caused by the group of bacteria under investigation, the most successful regimen was monotherapy with carbapenems (doripenem, meropenem and imipenem). Cephalosporins (cefepim and ceftazidim) were less effective in vitro. The worst results were obtained with the combination of piperacillin/tazobactam with aminoglycosides (amikacin or gentamicin) or fluoroquinolones (ciprofloxacin). CONCLUSIONS: Antibiotic monotherapy proved to be more effective in VAP patients than combined therapy; the best results were achieved with carbapenems. Doripenem showed strong activity in vitro against P. aeruginosa and Klebsiella sp. and should be considered for empirical VAP therapy; however, carbapenems may be less effective against Acinetobacter baumannii. The wide range of bacteria, and their broad range of susceptibility to antibiotics, suggests the need for modification of current recommendations.

The phenotypic and genetic biofilm formation characteristics of coagulase-negative staphylococci isolates in children with otitis media.

OBJECTIVE: Medical biofilms are involved in a number of chronic infections including otitis media with effusion and chronic rhinosinusitis, which are common pediatric infectious diseases. The purpose of the study was to analyze the phenotypic and genotypic indicators of biofilm formation of coagulase negative staphylococci isolates in children with otitis media with effusion, and in children with chronic rhinosinusitis as a comparison group by using three different detection methods. METHODS: Forty nine children aged from 2 to 6 years old, diagnosed with otitis media with effusion were enrolled to the study. The comparative group consisted of twenty three strains of coagulase-negative staphylococci from the strains collection isolated from nose swabs from children 3 to 7 years old suffering from rhinosinusitis for longer than 12 weeks. Cultured strains were tested for biofilm formation ability with three tests: Congo red agar, tissue culture plate methods and detection of ica operon. RESULTS: Out of 97 ear effusion specimens, obtained from 49 children suffering from OME, 38 were found positive in conventional culture resulting in isolation of 50 different bacterial species. Nested-PCR method confirmed bacterial presence in 95 (97.9%) cases. Among 50 different bacterial species isolated, 30 (30.9%) CNS and 20 (20.6%) other than CNS species. Detection of slime producing phenotype of CNS was performed with CRA plate test. Among OME isolates, 11 (36.7%) were CRA plate test positive. In case of isolates from CRS, 8 (34.8%) strains revealed black coloration on CRA. Using TCP method, strong adherence to microtiter plate was observed in two Staphylococcus epidermidis strains from OME and two S. epidermidis from CRS. By using the ica operon test, the genotypic ability to form biofilm was identified in 7 (23.3%) S. epidermidis strains cultured from ears effusion and in 3 (13%) strains from nose swabs. CONCLUSIONS: CNS strains revealed genotypic and phenotypic features responsible for the ability to form the biofilm in vivo. The presence of ica genes and phenotypic ability to form a biofilm by CNS strains emphasizes the pathogenic character of these strains in some cases of otitis media with effusion.

Intestinal colonization with Oxalobacter formigenes and its relation to urinary oxalate excretion in pediatric patients with idiopathic calcium urolithiasis.

BACKGROUND AND AIM: Oxalobacter formigenes is an intestinal bacterium that utilizes oxalate as the only source of energy. It has been suggested that the lack of colonization with this organism may be a risk factor for calcium oxalate urolithiasis. Because this problem was not investigated in pediatric stone formers, we decided to assess it in our patients. METHODS: The presence of O. formigenes in stool samples of 76 children and adolescents (aged 4.1-18 years) with idiopathic calcium urolithiasis (36 with chemically confirmed calcium oxalate stones and 40 children with a strong clinical suspicion of this type of urolithiasis) was assessed using PCR method. Simultaneously, urinary oxalate excretion was measured in this group. Fifty healthy, age- and sex-matched subjects served as controls. RESULTS: O. formigenes was found in 21/76 patients (27.6%). In controls, frequency of colonization was similar (26%). The median 24h urinary oxalate excretion in patients colonized with O. formigenes was significantly lower in comparison with non-colonized patients, 0.319 (range 0.141-0.546) and 0.437 (range 0.198-0.967) mmol/1.73 m(2)/24h, respectively. CONCLUSIONS: Higher urinary oxalate excretion in children with calcium urolithiasis may be a result of the absence of O. formigenes. The reasons for similarly low intestinal colonization with this bacterium in normal subjects and stone formers remain speculative. Thus, further studies are necessary to clarify this issue.

Covalent coating of hydroxyapatite by keratin stabilizes gentamicin release.

A novel hybrid hydroxyapatite (HAP) matrix, covalently coated with rarely applied, hardly degradable keratin and effectively modified by gentamicin immobilized in mixed-type mode (via interactions of diverse strength), was created. This hybrid showed a remarkably high drug immobilization yield and the most sustainable antibiotic release among all tested composites. It was also able to inhibit bacterial growth, both in surrounding liquid and on matrix surface, much longer (for at least 121 days of experiment) than analogous gelatin-modified and nonmodified matrices. Gentamicin-keratin-coated-HAP granules were nontoxic to human osteoblasts and enabled their proliferation with a rate similar as noncoated HAP. Presence of keratin on HAP granules seemed to slightly enhance the osteoblast proliferation. The results indicate that newly created HAP hybrid with covalently immobilized keratin and gentamicin--nontoxic and osteoblast-friendly--is a promising biomaterial of significantly prolonged antibacterial activity.

Monitoring Legionella species in hospital water systems. Link with disease and evaluation of different detection methods.

The aim of this work was to evaluate three currently available isolation methods for Legionella using water samples and swabs of a single pediatric hospital water system. Additionally, high risk patients were screened for the presence of Legionella pneumophila antigen in urine. Fifteen water samples and 11 swab samples were collected from distal sites at 18 sampling locations. The International Standard Method (PN-ISO11731-2) based on membrane filtration and direct culture of bacteria on selective media were compared with amoebic co-culture. The numbers of legionellae detected exceeded 10(2) cfu/100 ml in 50% of the samples. All the positive samples contained L. pneumophila SGs 2-14. Urine samples were obtained from 57 immunosuppressed children and screened for the presence of L. pneumophila serogroup (SG) 1 antigen by Legionella urinary antigen EIA. Of the 57 urine samples tested for the presence of Legionella pneumophila SG 1 antigen, none were positive. Our results highlight the value of combined membrane filtration and amoebic co-culture methods in detecting viable L. pneumophila strains. Direct plating of 0.2 ml water is a useful screening method for samples containing large bacterial amount.

[Etiological factors of vascular grafts infections and their resistance to antibiotics]. / Czynniki etiologiczne zakazen przeszczepów naczyniowych i ocena ich lekoopornosci.

THE AIM: of our study was the identification of microorganisms causing vascular graft infections and the evaluation of their antimicrobial susceptibility. MATERIAL AND METHODS: 25 patients with infected vascular graft, took part in our research. In 19 patients late type of infection was recognized, in 6 the infection was qualified as early. Purulent discharge obtained from the fistula was inoculated on the bacteriological media. Antimicrobial susceptibility was assessed by disc-diffusion method. RESULTS: Staphylococcus aureus and Pseudomonas aeruginosa proved to be the most frequently isolated microorganisms. Mixed infection, caused by two distinct bacteria, occurred in 5 patients; in all cases one species belonged to Gram-positive, and the second one to Gram-negative bacteria. In 50% of patients with early type infection different species of Gram-negative rods were present, in 37,5% of them S. aureus and S. epidermidis were isolated. In late type infection Gram-negative rods were isolated from 54,5% of patients and Gram-positive bacteria from 31,5% of patients. The most frequently isolated species appeared to be Pseudomonas aeruginosa. The isolated species of bacteria varied depending on the degree of infection (according to Shilagy and Samson). CONCLUSIONS: A diversity of isolated species, the presence of mixed infections and resistance patterns typical for hospital flora among bacteria infecting vascular grafts cause that antibiotic therapy should always be based on the results of microbiological examination.

Chlamydia pneumoniae infection in abdominal aortic aneurysm (AAA) patients and its clinical impact.

The aim of our study was to assess the presence of Chlmydia pneumoniae infection in AAA patients and to evaluate its association with clinical symptoms and histological signs of inflammation in the aortal wall. Fifty-two AAA patients participated in the research. Thirty healthy controls took part in serological examination. C. pneumoniae was detected by PCR and immunofluorescence in situ reaction in aorta samples of 84.6% and 86.54% of the patients, respectively. Serological markers of chronic C. pneumoniae infection were detected in 86.5% of AAA patients and in 33.3% of healthy controls. High titers of specific IgG and IgA were found in 37.8% of AAA patients with serologically defined chronic infection. All patients in "high serology" group had symptomatic aneurysm and inflammatory infiltrations in their aortal wall samples. AAA patients infected with C. pneumoniae are not a homogenous group. "High serology" group is much more prone to have symptomatic aneurysm than the remaining of AAA patients. Serology can be very useful in predicting the risk of AAA rupture. Inter-laboratory standardization of direct and indirect detection methods of C. pneumoniae infection is required to elucidate the role of these bacteria in AAA development.

Evaluation of the association between atypical bacteria infections and respiratory tract diseases with emphasis on bronchial asthma exacerbations in children.

Mycoplasma pneumoniae and Chlamydia pneumonie are important etiological agents responsible for human respiratory tract diseases. Recently, these atypical microorganisms received much attention regarding their role in bronchial asthma pathogenesis, which is one of the most frequent chronic diseases in children. The aim of the study was to investigate the association between infections caused by these pathogens and respiratory tract diseases in children. Levels of M. pneumoniae and C. pneumoniae-specific antibodies were determined in serum samples obtained from 30 patients suffering from bronchial asthma exacerbations, 10 patients with pneumonia, 28 patients with chronic otitis media with effusion (COME) and 22 sinusitis patients. Specific anti-M. pneumoniae antibodies were detected more frequently in the patients enrolled in the study than in control subjects. The highest percentage of the serum samples, which demonstrated the presence of M. pneumoniae-specific antibodies was demonstrated in patients with asthma (60%) and it was twofold higher than in control subjects. Serologic profile of 26.6% patients with asthma, 50% of patients with pneumonia, 39.2% of patients with COME, 45.4% of patients with sinusitis and 10% of control subjects was consistent with a possible acute infection caused by M. pneumoniae. The presence of specific anti-C. pneumoniae antibodies was demonstrated in a smaller percentage of patients--in 13.3% of children with asthma, 10% of children with pneumonia and in 7.1% of patients with COME; the level of specific antiobodies was suggestive of acute chlamydial infection only in COME patients. Analysis of serologic markers for atypical bacteria infections indicates a possible association between infections caused by M. pneumoniae and bronchial asthma exacerbations and other respiratory tract disorders including pneumonia, sinusitis and COME.

Evaluation of an anti-chlamydial antibiotic therapy influence on asthma patients.

Chlamydia pneumoniae is one of the most frequent pathogens causing airways infections. Contribution of chronic chlamydial infection to the following diseases: asthma, POChP, coronary heart disease, abdominal aortic aneurysm, is particularly interesting. The connection between such infection and bronchial asthma was described in the literature in 1991. C. pneumoniae often causes asthma exacerbation; it is suggested that it also may be an etiologic factor of the disease. In a group of 55 subjects with chronic, stable bronchial asthma treated in the Pulmonary Department, serologic characteristic of C. pneumoniae infection was found in 34 patients (61,8%). Thirteen of these subjects agreed to participate in the study. They were divided into two groups; placebo was administered to the first one and azithromycin in a dose of 1000 mg once a week--to the other one. The research was conducted using the double blind trial method. Anti-chlamydial antibody level was evaluated before and after treatment. Spirometry tests as well as subjective estimation of physical fitness and dyspnoea degree were also determined. In comparison with 'the placebo group', statistically significant improvement in respiratory parameters 'in the treated group' was not ascertained.

Immunofluorescence in situ and the serologic indices of Chlamydia pneumoniae infection in patients with an abdominal aortic aneurysm.

The aim of the study was to evaluate the frequency of Chlamydia pneumoniae infection in patients with abdominal aortic aneurysm (AAA) using selected methods. The histological specimens of aneurysm wall were evaluated, the method of immunofluorescence was used to reveal the antigen in the wall of AAA and the titers of specific antibodies of IgG, IgM and IgA classes in blood plasma were marked. Atherosclerotic changes in the aneurysm wall were found in all patients. In 20(87%) patients the C. pneumoniae antigen was seen in the wall of abdominal aneurysm using the indirect immunofluorescence method. A significant relation between the method of direct C. pneumoniae diagnosis, aneurysm symptoms and histologically detected inflammation in its wall was confirmed. Serologic markers of the chronic C. pneumoniae infection were seen in 20(87%) out of 23 patients and in 6(30%) out of 20 subjects of the control group and this difference was statistically significant. It was observed, that all patients with serologic indices of active C. pneumoniae infection, had symptomatic aneurysm. The presence of Chlamydia pneumoniae in the wall of AAA as well as the occurrence of serologic indices of the chronic infection in these patients can confirm the hypothesis of the relation between the infection with this microorganism and the development of the disease.