Burkholderia stagnalis sp. nov. and Burkholderia territorii sp. nov., two novel Burkholderia cepacia complex species from environmental and human sources.

Nine Burkholderia cepacia complex (Bcc) bacteria were isolated during environmental surveys for the ecological niche of Burkholderia pseudomallei, the aetiological agent of melioidosis, in the Northern Territory of Australia. They represented two multi-locus sequence analysis-based clusters, referred to as Bcc B and Bcc L. Three additional environmental and clinical Bcc B isolates were identified upon deposition of the sequences in the PubMLST database. Analysis of the concatenated nucleotide sequence divergence levels within both groups (1.4 and 1.9%, respectively) and towards established Bcc species (4.0 and 3.9%, respectively) demonstrated that the two taxa represented novel Bcc species. All 12 isolates were further characterized using 16S rRNA and recA gene sequence analysis, RAPD analysis, DNA base content determination, fatty acid methyl ester analysis and biochemical profiling. Analysis of recA gene sequences revealed a remarkable diversity within each of these taxa, but, together, the results supported the affiliation of the two taxa to the Bcc. Bcc B strains can be differentiated from most other Bcc members by the assimilation of maltose. Bcc L strains can be differentiated from other Bcc members by the absence of assimilation of N-acetylglucosamine. The names Burkholderia stagnalis sp. nov. with type strain LMG 28156(T) ( = CCUG 65686(T)) and Burkholderia territorii sp. nov. with type strain LMG 28158(T) ( = CCUG 65687(T)) are proposed for Bcc B and Bcc L bacteria, respectively.

Whole-genome sequencing confirms that Burkholderia pseudomallei multilocus sequence types common to both Cambodia and Australia are due to homoplasy.

Burkholderia pseudomallei isolates with shared multilocus sequence types (STs) have not been isolated from different continents. We identified two STs shared between Australia and Cambodia. Whole-genome analysis revealed substantial diversity within STs, correctly identified the Asian or Australian origin, and confirmed that these shared STs were due to homoplasy.

Bloodstream infection among adults in Phnom Penh, Cambodia: key pathogens and resistance patterns.

BACKGROUND: Bloodstream infections (BSI) cause important morbidity and mortality worldwide. In Cambodia, no surveillance data on BSI are available so far. METHODS: From all adults presenting with SIRS at Sihanouk Hospital Centre of HOPE (July 2007-December 2010), 20 ml blood was cultured. Isolates were identified using standard microbiological techniques; antibiotic susceptibilities were assessed using disk diffusion and MicroScan®, with additional E-test, D-test and double disk test where applicable, according to CLSI guidelines. RESULTS: A total of 5714 samples from 4833 adult patients yielded 501 clinically significant organisms (8.8%) of which 445 available for further analysis. The patients' median age was 45 years (range 15-99 y), 52.7% were women. HIV-infection and diabetes were present in 15.6% and 8.8% of patients respectively. The overall mortality was 22.5%. Key pathogens included Escherichia coli (n = 132; 29.7%), Salmonella spp. (n = 64; 14.4%), Burkholderia pseudomallei (n = 56; 12.6%) and Staphylococcus aureus (n = 53; 11.9%). Methicillin resistance was seen in 10/46 (21.7%) S. aureus; 4 of them were co-resistant to erythromycin, clindamycin, moxifloxacin and sulphamethoxazole-trimethoprim (SMX-TMP). We noted combined resistance to amoxicillin, SMX-TMP and ciprofloxacin in 81 E. coli isolates (62.3%); 62 isolates (47.7%) were confirmed as producers of extended spectrum beta-lactamase. Salmonella isolates displayed high rates of multidrug resistance (71.2%) with high rates of decreased ciprofloxacin susceptibility (90.0%) in Salmonella Typhi while carbapenem resistance was observed in 5.0% of 20 Acinetobacter sp. isolates. CONCLUSIONS: BSI in Cambodian adults is mainly caused by difficult-to-treat pathogens. These data urge for microbiological capacity building, nationwide surveillance and solid interventions to contain antibiotic resistance.

Azithromycin and ciprofloxacin resistance in Salmonella bloodstream infections in Cambodian adults.

BACKGROUND: Salmonella enterica is a frequent cause of bloodstream infection (BSI) in Asia but few data are available from Cambodia. We describe Salmonella BSI isolates recovered from patients presenting at Sihanouk Hospital Centre of Hope, Phnom Penh, Cambodia (July 2007-December 2010). METHODOLOGY: Blood was cultured as part of a microbiological prospective surveillance study. Identification of Salmonella isolates was performed by conventional methods and serotyping. Antibiotic susceptibilities were assessed using disk diffusion, MicroScan and E-test macromethod. Clonal relationships were assessed by Pulsed Field Gel Electrophoresis; PCR and sequencing for detection of mutations in Gyrase and Topoisomerase IV and presence of qnr genes. PRINCIPAL FINDINGS: Seventy-two Salmonella isolates grew from 58 patients (mean age 34.2 years, range 8-71). Twenty isolates were identified as Salmonella Typhi, 2 as Salmonella Paratyphi A, 37 as Salmonella Choleraesuis and 13 as other non-typhoid Salmonella spp. Infection with human immunodeficiency virus (HIV) was present in 21 of 24 (87.5%) patients with S. Choleraesuis BSI. Five patients (8.7%) had at least one recurrent infection, all with S. Choleraesuis; five patients died. Overall, multi drug resistance (i.e., co-resistance to ampicillin, sulphamethoxazole-trimethoprim and chloramphenicol) was high (42/59 isolates, 71.2%). S. Typhi displayed high rates of decreased ciprofloxacin susceptibility (18/20 isolates, 90.0%), while azithromycin resistance was very common in S. Choleraesuis (17/24 isolates, 70.8%). Two S. Choleraesuis isolates were extended spectrum beta-lactamase producer. CONCLUSIONS AND SIGNIFICANCE: Resistance rates in Salmonella spp. in Cambodia are alarming, in particular for azithromycin and ciprofloxacin. This warrants nationwide surveillance and revision of treatment guidelines.

Clara cell protein in bronchoalveolar lavage fluid: a predictor of ventilator-associated pneumonia?

INTRODUCTION: Clara cell protein 10 (CC-10) has been associated with inflammatory and infectious pulmonary diseases. This study evaluates CC-10 concentrations in bronchoalveolar lavage (BAL) fluid as a potential marker of ventilator-associated pneumonia (VAP). METHODS: Between January 2003 and December 2007, BAL fluid samples obtained from critically ill patients at the intensive care unit of the Maastricht University Medical Centre clinically suspected of having VAP were included. Patients were divided into two groups: (1) microbiologically confirmed VAP (the VAP group) and (2) microbiologically unconfirmed VAP (the non-VAP group). The concentration of CC-10 was measured by means of a commercially available enzyme-linked immunosorbent assay kit, and retrospective analysis was performed. Areas under the curve of receiver operating characteristic curves were calculated for CC-10 concentrations. RESULTS: A total of 196 patients (122 men, 74 women) were included. A total of 79 (40%) of 196 cases of suspected VAP were microbiologically confirmed. The median CC-10 concentration in the VAP group was 3,019 ng/mL (range, 282 to 65,546 ng/mL) versus 2,504 ng/mL (range, 62 to 30,240 ng/mL) in the non-VAP group (P = 0.03). There was no significant difference in CC-10 concentrations between patients treated with or without corticosteroids (P = 0.26) or antibiotic therapy (P = 0.9). The CC-10 concentration did not differ significantly between patients with Gram-positive versus Gram-negative bacteria that caused the VAP (P = 0.06). However, CC-10 concentrations did differ significantly between the late-onset VAP group and the non-VAP group. CONCLUSIONS: The CC-10 concentration in BAL fluid yielded low diagnostic accuracy in confirming the presence of VAP.

Trends of norfloxacin and erythromycin resistance of Campylobacter jejuni/Campylobacter coli isolates recovered from international travelers, 1994 to 2006.

BACKGROUND: Campylobacter sp. is a major cause of bacterial enterocolitis and travelers' diarrhea. Empiric treatment regimens include fluoroquinolones and macrolides. METHODS: Over the period 1994 to 2006, 724 Campylobacter jejuni/Campylobacter coli isolates recovered from international travelers at the outpatient clinic of the Institute of Tropical Medicine, Antwerp, Belgium, were reviewed for their susceptibility to norfloxacin and erythromycin. RESULTS: Norfloxacin resistance increased significantly over time in isolates from travelers returning from Asia, Africa, and Latin America. For the years 2001 to 2006, norfloxacin resistance rates were 67 (70.5%) of 95 for Asia, 20 (60.6%) of 33 for Latin America, and 36 (30.6%) of 114 for Africa. The sharpest increase was noted for India, with no resistance in 1994, but 41 (78.8%) of 52 resistant isolates found during 2001 to 2006. Erythromycin resistance was demonstrated in 20 (2.7%) isolates, with a mean annual resistance of 3.1% +/- 2.8%; resistance increased over time, with up to 3(7.5%) of 40 and 3 (8.6%) of 35 resistant isolates in 2004 and 2006, respectively (p < 0.05); there was no apparent geographic association. Combined resistance to norfloxacin and erythromycin was observed in five isolates. CONCLUSIONS: The high resistance rates to fluoroquinolones warrant reconsideration of their use as drugs of choice in patients with severe gastroenteritis when Campylobacter is the presumed cause. Continued monitoring of the incidence and the spread of resistant Campylobacter isolates is warranted.

Biometric fingerprinting for visa application: device and procedure are risk factors for infection transmission.

BACKGROUND: Biometric fingerprint identity verification is currently introduced in visa application and entry screening at border control. The system implies physical contact between the skin and the surface of the fingerprint-capturing and reading devices. AIM: To assess the risk of infection transmission through fingerprinting. METHODS: The medical literature was reviewed for the potential of microorganisms to be carried on the skin of hands in the community, to be transferred from hands to inanimate surfaces, to survive on surfaces, and to be transferred in doses exceeding the infectious dose. The fingerprinting procedures as currently applied were reviewed. RESULTS: Factors that favor transfer of microorganisms are large skin-surface contact between flat fingers (2 x 20 cm(2)) and fingerprint-capturing device, nonporous contact surface, large overlap of contact surface and short turnaround time between successive applicants, high contact pressure, and difficulties to disinfect devices. Transmission risk exists for enteric viruses (rotavirus, norovirus, and hepatitis A virus), respiratory viruses (respiratory syncytial virus, rhinovirus, influenza virus, etc.), and enteropathogenic bacteria with low infectious doses (Shigella dysenteriae, Enterohemorrhagic Escherichia coli, etc.). Using Monte Carlo risk analysis on US data, transmission of human rotavirus is estimated at 191 [95% credible intervals (CI) 0-289] per million fingerprint-capturing procedures. Application of 70% isopropyl hand rub and 85% ethanol hand gel reduces the risk to 77 (95% CI 0-118) and 0.3 (95% CI 0-0.3) transmissions per million procedures, respectively. CONCLUSIONS: The fingerprinting procedure as currently used is associated with a risk of infection transmission. Simple hygienic measures can considerably reduce this transmission risk.

Herpes simplex virus load in bronchoalveolar lavage fluid is related to poor outcome in critically ill patients.

OBJECTIVE: To evaluate the relationship between the HSV-1 and -2 loads in BAL fluid (BALF) and clinical outcome. DESIGN: Retrospective study. SETTING: The general intensive care unit of the University Hospital Maastricht. PATIENTS: Five hundred and twenty-one BALF samples from 462 patients were included. Patients were divided into three groups; (1) patients admitted to the hospital <48 h before lavage (Community), (2) patients admitted to the ICU >48 h before lavage (ICU) and (3) the remaining patients (non-ICU group). INTERVENTIONS: No additional interventions were conducted. MEASUREMENTS AND RESULTS: HSV-1 and HSV-2 loads were determined by real-time polymerase chain reaction (PCR). HSV-1 DNA was detected in 4.3% (4/92) of samples in the community group, 15% (18/121) in the non-ICU group and in 32% (99/308) of the ICU group. In the age group <50 years HSV-1 DNA was less frequently isolated compared to the age group >or=50 years (16/129 (12%) versus 187/376 (25%), respectively, OR = 2.6; P < 0.001). HSV-1 loads of >10(5) genome equivalents (ge)/ml were associated with an increased 14-day in-hospital mortality compared to patients with a HSV-1 load 10(5) ge/ml enforces its clinical relevance and necessitates to start randomized medical intervention studies.

Rapid and reliable identification of Streptococcus anginosus group isolates to the species level by real-time PCR and melting curve analysis.

A real-time PCR assay is described for rapid and accurate identification of the Streptococcus anginosus group members. Based on melting curve analysis, the S. anginosus species isolates could be further subdivided into two subgroups, each associated with different clinically relevant ribotypes of S. anginosus.

C-reactive protein and procalcitonin concentrations in bronchoalveolar lavage fluid as a predictor of ventilator-associated pneumonia.

BACKGROUND: Diagnosis of ventilator-associated pneumonia (VAP) is difficult. The usefulness of high-sensitivity procalcitonin (ProCa-S) and high-sensitivity C-reactive protein (CRPH) in bronchoalveolar lavage (BAL) fluid and serum in the prediction of VAP was determined. METHODS: The study was conducted over a 28-month period (November 1999-June 2002) at the University Hospital Maastricht. BAL fluid samples were collected from patients admitted to the intensive care unit. Differential cell count and quantitative culture of BAL fluid were performed. C-reactive protein (CRP) and procalcitonin (PCT) on BAL fluid were determined by means of two high-sensitivity kits (CRPH and ProCa-S, respectively). Since both kits were designed for use on serum, validation for use on BAL fluid was performed. RESULTS: A total of 117 patients were included. 43.6% (51/117) had microbiologically confirmed VAP. Both CRPH and ProCa-S showed good matrix effect, linearity and intra- and inter-assay variation. No significant differences in PCT and CRP concentrations in serum and BAL fluid were found between the VAP and the non-VAP group. CONCLUSIONS: Both the ProCa-S and the CRPH kits can be used for assessing the concentration of PCT and CRP in BAL fluid, respectively. PCT and CRP concentrations in BAL fluid appeared to be of no additional value in the diagnosis of VAP.

Influence of antibiotic therapy on the cytological diagnosis of ventilator-associated pneumonia.

OBJECTIVE: To assess the influence of antibiotics on the value of various cytological parameters, and their combinations, in diagnosing ventilator-associated pneumonia (VAP). DESIGN: Prospective study. SETTING: The general intensive care unit (17 beds) of the University Hospital Maastricht. PATIENTS: Three hundred and thirty-five episodes of clinically suspected VAP (defined by the clinical and radiological criteria previously described by Bonten et al.) in 282 patients were studied. INTERVENTIONS: No additional interventions were conducted. MEASUREMENTS AND RESULTS: Bronchoalveolar lavage fluid cytology included a total cell count per millilitre, differential cell count and the percentage of infected cells (cells containing phagocytised organisms). Antibiotic therapy from 72 h prior to lavage was recorded. Areas under the curve (AUCs) of receiver operating characteristic curves were calculated for various cytological parameters and their combinations, in patients with and without antibiotic therapy. In 126 episodes (37.6%) in 106 patients, VAP was confirmed. There was no difference in AUCs between patients with and without antibiotic therapy for any parameter studied. The most prominent AUCs were (for patient groups with and without antibiotics combined): total cell count, 0.65; percentage polymorphonuclear neutrophils, 0.71; and percentage infected cells, 0.90. The combination of percentage infected cells with any other cytological parameter did not increase the AUC. CONCLUSION: Antibiotic therapy did not influence the predictive value of the percentage infected cells in BALF in diagnosing VAP.

Exposure to bacterial DNA before hemorrhagic shock strongly aggravates systemic inflammation and gut barrier loss via an IFN-gamma-dependent route.

OBJECTIVE: To investigate the role of bacterial DNA in development of an excessive inflammatory response and loss of gut barrier loss following systemic hypotension. SUMMARY BACKGROUND DATA: Bacterial infection may contribute to development of inflammatory complications following major surgery; however, the pathogenesis is not clear. A common denominator of bacterial infection is bacterial DNA characterized by unmethylated CpG motifs. Recently, it has been shown that bacterial DNA or synthetic oligodeoxynucleotides containing unmethylated CpG motifs (CpG-ODN) are immunostimulatory leading to release of inflammatory mediators. METHODS: Rats were exposed to CpG-ODN prior to a nonlethal hemorrhagic shock. The role of interferon-gamma (IFN-gamma) was investigated by administration of anti IFN-gamma antibodies. RESULTS: Exposure to CpG-ODN prior to hemorrhagic shock significantly augmented shock-induced release of IFN-gamma, tumor necrosis factor-alpha (TNF-alpha) (P < 0.05), interleukin (IL)-6 (P < 0.05), and nitrite levels (P < 0.05), while there was a defective IL-10 response (P < 0.05). Simultaneously, expression of Toll-like receptor (TLR) 4 in the liver was markedly enhanced. Furthermore, intestinal permeability for HRP significantly increased and bacterial translocation was enhanced in hemorrhagic shock rats pretreated with CpG-ODN. Interestingly, inhibition of IFN-gamma in CpG-treated animals reduced TNF-alpha (P < 0.05), IL-6 (P < 0.05), nitrite (P < 0.05), and intestinal permeability following hemorrhagic shock (P < 0.05) and down-regulated expression of TLR4. CONCLUSION: Exposure to bacterial DNA strongly aggravates the inflammatory response, disrupts the intestinal barrier, and up-regulates TLR4 expression in the liver following hemorrhagic shock. These effects are mediated via an IFN-gamma-dependent route. In the clinical setting, bacterial DNA may be important in development of inflammatory complications in surgical patients with bacterial infection.

Inter-laboratory comparison of three different real-time PCR assays for the detection of Pneumocystis jiroveci in bronchoalveolar lavage fluid samples.

Pneumocystis jiroveci pneumonia (PCP) is an opportunistic infection affecting immunocompromised patients. While conventional diagnosis of PCP by microscopy is cumbersome, the use of PCR to diagnose PCP has great potential. Nevertheless, inter-laboratory validation and standardization of PCR assays is lacking. The aim of this study was to evaluate the inter-laboratory agreement of three independently developed real-time PCR assays for the detection of P. jiroveci in bronchoalveolar lavage fluid samples. Therefore, 124 samples were collected in three tertiary care laboratories (Leiden University Medical Center, Maastricht Infection Center and Radboud University Nijmegen Medical Centre) and were tested by both microscopy and real-time PCR. Of 41 samples positive for P. jiroveci by microscopy, 40 were positive in all three PCR assays. The remaining sample was positive in a single assay only. Out of 83 microscopy-negative samples, 69 were negative in all three PCR assays. The other 14 samples were found positive, either in all three assays (n=5), in two (n=2) or in one of the assays (n=7). The data demonstrate high inter-laboratory agreement among real-time PCR assays for the detection of P. jiroveci.

Student perceptions about the characteristics of an effective discussion during the reporting phase in problem-based learning.

To explore student perceptions of factors contributing to the effectiveness of discussions in the reporting phase of the problem-based learning (PBL) process, where students report and synthesise the results of self-study. Forty-eight Year 1 and 2 medical students participated in 6 focus group interviews about the characteristics of effective group discussions and possible improvements. The data were analysed qualitatively in several stages. The analysis yielded 4 main characteristics of effective discussions: asking for, giving and receiving explanations; integrating and applying knowledge; discussing differences with regard to learning content, and guiding and monitoring the content and the group process of the discussion. Integrating and applying knowledge included structuring, relating and summarising information and providing examples from practice. Discussing different opinions included discussing a variety of literature resources and disagreements. The main learning effects mentioned by the students were retention, understanding, integration and application of knowledge. Students have clear ideas about what promotes effective discussions during the reporting phase. Their PBL experience has provided them with some insights that are in line with theory and research on collaborative learning. Future research should examine differences between student and tutor perceptions of the quality of discussions. Introductions to PBL for students and tutors should include training in asking open but focused questions, supporting explanations with arguments and dealing with conflicts about learning content. Tutors should be trained in giving effective and personal feedback. Collaborative creation of external knowledge representations (i.e. concept maps) should be advocated, as should variety of literature resources.

Patterns of inflammation and the use of reversibility testing in smokers with airway complaints.

BACKGROUND: Although both smoking and respiratory complaints are very common, tools to improve diagnostic accuracy are scarce in primary care. This study aimed to reveal what inflammatory patterns prevail in clinically established diagnosis groups, and what factors are associated with eosinophilia. METHOD: Induced sputum and blood plasma of 59 primary care patients with COPD (n = 17), asthma (n = 11), chronic bronchitis (CB, n = 14) and smokers with no respiratory complaints ('healthy smokers', n = 17) were collected, as well as lung function, smoking history and clinical work-up. Patterns of inflammatory markers per clinical diagnosis and factors associated with eosinophilia were analyzed by multiple regression analyses, the differences expressed in odds ratios (OR) with 95% confidence intervals. RESULTS: Multivariately, COPD was significantly associated with raised plasma-LBP (OR 1.2 [1.04-1.37]) and sTNF-R55 in sputum (OR 1.01 [1.001-1.01]), while HS expressed significantly lowered plasma-LBP (OR 0.8 [0.72-0.95]). Asthma was characterized by higher sputum eosinophilic counts (OR 1.3 [1.05-1.54]), while CB showed a significantly higher proportion of sputum lymphocytic counts (OR 1.5 [1.12-1.9]). Sputum eosinophilia was significantly associated with reversibility after adjusting for smoking, lung function, age, gender and allergy. CONCLUSION: Patterns of inflammatory markers in a panel of blood plasma and sputum cells and mediators were discernable in clinical diagnosis groups of respiratory disease. COPD and so-called healthy smokers showed consistent opposite associations with plasma LBP, while chronic bronchitics showed relatively predominant lymphocytic inflammation compared to other diagnosis groups. Only sputum eosinophilia remained significantly associated with reversibility across the spectrum of respiratory disease in smokers with airway complaints.

Rapid identification of bacteria by real-time amplification and sequencing of the 16S rRNA gene.

To allow rapid identification of bacteria in pure cultures and blood culture bottles, an assay was developed which is based on real-time amplification and sequencing of bacterial 16 S rRNA genes. In principle, this assay allows identification of bacteria from pure cultures within 6.5 h, and from blood cultures within approximately 7 h.

Case studies to explore the pitfalls in the diagnosis of sarcoidosis.

Sarcoidosis is a multisystemic disorder of unknown cause. Diagnosis is established when clinical and radiological findings are supported by histological evidence of noncaseating epitheloid cell granulomas. Exclusion of granulomas of known causes and sarcoidlike reactions is mandatory. A lot of infections may mimic a sarcoidlike granulomatous reaction. Even with well advanced pathological and microbiological examination, it could be hard to make the appropriate diagnosis. Moreover, sarcoidosis patients receiving corticosteroids are susceptible to opportunistic infections. The challenge is, however, making the right diagnosis because opportunistic infections can resemble sarcoidosis. The case reports presented in this paper are meant to stress the importance of excluding granulomatous infections in patients with (suspected) sarcoidosis. Appropriate diagnostic procedures are important to exclude an infectious condition mimicking sarcoidosis. Accordingly, appropriate treatment can start without further delay.

Nutritional stimulation of cholecystokinin receptors inhibits inflammation via the vagus nerve.

The immune system in vertebrates senses exogenous and endogenous danger signals by way of complex cellular and humoral processes, and responds with an inflammatory reaction to combat putative attacks. A strong protective immunity is imperative to prevent invasion of pathogens; however, equivalent responses to commensal flora and dietary components in the intestine have to be avoided. The autonomic nervous system plays an important role in sensing luminal contents in the gut by way of hard-wired connections and chemical messengers, such as cholecystokinin (CCK). Here, we report that ingestion of dietary fat stimulates CCK receptors, and leads to attenuation of the inflammatory response by way of the efferent vagus nerve and nicotinic receptors. Vagotomy and administration of antagonists for CCK and nicotinic receptors significantly blunted the inhibitory effect of high-fat enteral nutrition on hemorrhagic shock-induced tumor necrosis factor-alpha and interleukin-6 release (P < 0.05). Furthermore, the protective effect of high-fat enteral nutrition on inflammation-induced intestinal permeability was abrogated by vagotomy and administration of antagonists for CCK and nicotinic receptors. These data reveal a novel neuroimmunologic pathway, controlled by nutrition, that may help to explain the intestinal hyporesponsiveness to dietary antigens, and shed new light on the functionality of nutrition.

Detection of Streptococcus pneumoniae antigen in bronchoalveolar lavage fluid samples by a rapid immunochromatographic membrane assay.

We conducted a retrospective study to evaluate an immunochromatographic membrane test (ICT), applied to bronchoalveolar lavage (BAL) fluid samples obtained in patients with suspected pneumonia, for the detection of Streptococcus pneumoniae antigen. The NOW Streptococcus pneumoniae test was assessed on 96 BAL fluid samples. Sensitivity was tested in 20 samples obtained from patients diagnosed as having pneumococcal pneumonia (growth of S. pneumoniae in blood cultures and/or in BAL fluid samples of > or =10(4) CFU/ml). Specificity was tested in BAL fluid samples of nonpneumococcal etiology (n = 41) and in samples with no respiratory pathogen and a total bacterial count of <10(4) CFU/ml (n = 35). Using the ICT, pneumococcal antigen was detected in 29 (30.2%) BAL fluid samples, with a sensitivity of 95.0% (95% confidence interval [CI], 90.6% to 99.4%) and a specificity of 86.8% (95% CI, 80.1% to 93.8%). The ICT was easy to perform and revealed unequivocal and reproducible results. No interference was observed with high cell counts, red blood cells, or elevated protein levels. Four out of 10 false-positive readings occurred in samples with S. pneumoniae counts below the 10(4) CFU/ml threshold limit of pneumonia. In BAL fluid samples obtained after pneumococcal bacteremia, positive test results were found for up to 35 days after bacteremia. The ICT test applied to BAL fluid specimens is reproducible and accurate in the diagnosis of pneumococcal antigen. Further studies are required to establish the impact of the ICT on patient care.