Accuracy of real-time PCR, Gram stain and culture for Streptococcus pneumoniae, Neisseria meningitidis and Haemophilus influenzae meningitis diagnosis.

BACKGROUND: Although cerebrospinal fluid (CSF) culture is the diagnostic reference standard for bacterial meningitis, its sensitivity is limited, particularly when antibiotics were previously administered. CSF Gram staining and real-time PCR are theoretically less affected by antibiotics; however, it is difficult to evaluate these tests with an imperfect reference standard. METHODS AND FINDINGS: CSF from patients with suspected meningitis from Salvador, Brazil were tested with culture, Gram stain, and real-time PCR using S. pneumoniae, N. meningitidis, and H. influenzae specific primers and probes. An antibiotic detection disk bioassay was used to test for the presence of antibiotic activity in CSF. The diagnostic accuracy of tests were evaluated using multiple methods, including direct evaluation of Gram stain and real-time PCR against CSF culture, evaluation of real-time PCR against a composite reference standard, and latent class analysis modeling to evaluate all three tests simultaneously. RESULTS: Among 451 CSF specimens, 80 (17.7%) had culture isolation of one of the three pathogens (40 S. pneumoniae, 36 N. meningitidis, and 4 H. influenzae), and 113 (25.1%) were real-time PCR positive (51 S. pneumoniae, 57 N. meningitidis, and 5 H. influenzae). Compared to culture, real-time PCR sensitivity and specificity were 95.0% and 90.0%, respectively. In a latent class analysis model, the sensitivity and specificity estimates were: culture, 81.3% and 99.7%; Gram stain, 98.2% and 98.7%; and real-time PCR, 95.7% and 94.3%, respectively. Gram stain and real-time PCR sensitivity did not change significantly when there was antibiotic activity in the CSF. CONCLUSION: Real-time PCR and Gram stain were highly accurate in diagnosing meningitis caused by S. pneumoniae, N. meningitidis, and H. influenzae, though there were few cases of H. influenzae. Furthermore, real-time PCR and Gram staining were less affected by antibiotic presence and might be useful when antibiotics were previously administered. Gram staining, which is inexpensive and commonly available, should be encouraged in all clinical settings.

Clinical outcome of pneumococcal meningitis during the emergence of pencillin-resistant Streptococcus pneumoniae: an observational study.

BACKGROUND: Prior to the availability of generic third-generation cephalosporins, penicillins were widely used for treatment of pneumococcal meningitis in developing countries despite concerns about rising levels of penicillin resistance among pneumococcal isolates. We examined the impact of penicillin resistance on outcomes of pneumococcal meningitis over a ten year period in an infectious diseases hospital in Brazil. METHODS: Clinical presentation, antimicrobial therapy and outcomes were reviewed for 548 patients with culture-confirmed pneumococcal meningitis from December, 1995, to November, 2005. Pneumococcal isolates from meningitis patients were defined as penicillin-resistant if Minimum Inhibitory Concentrations for penicillin were greater than 0.06 µg/ml. Proportional hazards regression was used to identify risk factors for fatal outcomes. RESULTS: During the ten-year period, ceftriaxone replaced ampicillin as first-line therapy for suspected bacterial meningitis. In hospital case-fatality for pneumococcal meningitis was 37%. Of 548 pneumococcal isolates from meningitis cases, 92 (17%) were resistant to penicillin. After controlling for age and severity of disease at admission, penicillin resistance was associated with higher case-fatality (Hazard Ratio [HR], 1.62; 95% Confidence Interval [CI], 1.08-2.43). Penicillin-resistance remained associated with higher case-fatality when initial therapy included ceftriaxone (HR, 1.68; 95% CI 1.02-2.76). CONCLUSIONS: Findings support the use of third generation cephalosporin antibiotics for treatment of suspected pneumococcal meningitis even at low prevalence of pneumococcal resistance to penicillins.

Serotype distribution and antimicrobial resistance of Streptococcus pneumoniae prior to introduction of the 10-valent pneumococcal conjugate vaccine in Brazil, 2000-2007.

This study describes the serotype distribution and antibiotic resistance patterns among 397 S. pneumoniae meningitis case isolates recovered in Salvador, Brazil, during the period of 2000-2007, before introduction of the 10-valent pneumococcal conjugate vaccine. The active hospital-based surveillance showed a decline in the annual incidence rates of pneumococcal meningitis during the period of study, from 1.12 cases to 0.83 cases/100,000 persons for all age groups (P<0.001), with an overall case-fatality rate of 28.6% (113 of 395) for all patients and 41.9% (57 of 136) for those <5 years of age. Serotypes 14 (n=55; 13.9%), 3 (n=32; 8.1%), 23F (n=32; 8.1%), 19F (n=31; 7.8%), 6B (n=30; 7.6%), 18C (n=28; 7.1%), and 6A (n=20; 5%) were the most prevalent serotypes. In patients <5 years the estimated projected coverage of 7-, 10- and 13-valent conjugate vaccines was 74.3%, 75.7% and 83.1%, respectively. Antimicrobial susceptibility testing revealed that 22.1% (n=88) of isolates were non-susceptible to penicillin, 56% were non-susceptible to trimethoprim/sulphamethoxazole, and 29.6% were non-susceptible to tetracycline. Nonsusceptibility to penicillin and cefotaxime was detected solely among serotype 14 isolates (n=4; 1%). This study provides an important baseline to assess the impact of conjugate vaccine implantation on the epidemiology of meningitis due to Streptococcus pneumoniae in Salvador, Brazil.

Poor clinical outcome for meningitis caused by Haemophilus influenzae serotype A strains containing the IS1016-bexA deletion.

Since the introduction of Haemophilus influenzae type b (Hib) conjugate vaccines, meningitis caused by serotypes other than Hib has gained in importance. We conducted active hospital-based surveillance for meningitis over an 11-year period in Salvador, Brazil. H. influenzae isolates were serotyped and analyzed by polymerase chain reaction, pulsed-field gel electrophoresis, and DNA sequencing to identify strains with a specific deletion (IS1016) in the bexA gene (IS1016-bexA). We identified 43 meningitis cases caused by non-type b H. influenzae: 28 (65%) were caused by type a (Hia), 9 (21%) were caused by noncapsulated strains, and 3 (7%) each were caused by types e and f. Hia isolates clustered in 2 clonal groups; clonal group A strains (n = 9) had the IS1016-bexA deletion. Among children <5 years of age, meningitis caused by Hia from clonal group A had higher case-fatality than meningitis caused by clonal group B. Despite small numbers, these results indicate that the presence of the IS1016-bexA deletion is associated with enhanced virulence in non-type b H. influenzae.

Insulin-like growth factor-I induced and constitutive arginase activity differs among isolates of Leishmania derived from patients with diverse clinical forms of Leishmania braziliensis infection.

Arginase activity has been related to leishmaniasis development, thus we studied the constitutive and insulin-like growth factor (IGF) I-induced arginase activity of Leishmania (Viannia) braziliensis isolates from patients with different clinical forms of American tegumentary leishmaniasis (ATL). Isolates from mucosal leishmaniasis presented higher basal levels of arginase activity than isolates from other clinical forms of ATL. Isolates from disseminated leishmaniasis that present mucosal lesion in some cases reached the arginase activity similar to that of isolates from mucosal leishmaniasis upon IGF-I stimulation. Differences in arginase activity may influence disease outcomes such as evolution to mucosal lesion in patients with L. (V.) braziliensis infection.

Burden of group A streptococcal meningitis in Salvador, Brazil: report of 11 years of population-based surveillance.

BACKGROUND: Over recent decades, a resurgence of invasive group A streptococcal (GAS) infections has been observed; GAS remains a rare cause of pyogenic meningitis. We report herein population-based findings of long-term surveillance for GAS meningitis in Salvador, Brazil, and estimate the overall burden of invasive GAS infections. METHODS: From February 1996 to February 2007 we conducted active surveillance for GAS meningitis in the state reference hospital for infectious diseases in Salvador, Brazil. Data on clinical presentation, laboratory records, and outcome were collected through interviews and chart review. GAS isolates were evaluated for antimicrobial susceptibility and emm type. RESULTS: We identified 20 cases of GAS meningitis, which accounted for 0.9% of all culture-proven bacterial meningitis in the study period. The mean annual incidence of GAS meningitis was 0.03 cases per 100,000 population in metropolitan Salvador and peaked in children <1 year of age (0.67 cases per 100,000 population). Among 17 cases with clinical information available, 41% required intensive care unit support and 25% died. Tested isolates were susceptible to penicillin and exhibited large emm type diversity. Based on the incidence of GAS meningitis, we estimate that the annual incidence of GAS infection is 3 cases per 100,000 population in metropolitan Salvador. CONCLUSIONS: Although rare, GAS is a life-threatening cause of bacterial meningitis. Knowledge of the incidence and emm type variability of the disease is necessary for planning immunization strategies.

Leptospirosis-associated severe pulmonary hemorrhagic syndrome, Salvador, Brazil.

We report the emergence of leptospirosis-associated severe pulmonary hemorrhagic syndrome (SPHS) in slum communities in Salvador, Brazil. Although active surveillance did not identify SPHS before 2003, 47 cases were identified from 2003 through 2005; the case-fatality rate was 74%. By 2005, SPHS caused 55% of the deaths due to leptospirosis.

Hospital-based surveillance of meningococcal meningitis in Salvador, Brazil.

This study aimed to describe the clinical, epidemiological and microbiological features of meningococcal meningitis in Salvador, Brazil. Between February 1996 and January 2001, a hospital-based surveillance prospectively identified cases of culture-positive meningococcal meningitis. Demographic and clinical data were collected through interview and medical chart review. Antisera and monoclonal antibodies were used to determine the serogroup and serotype:serosubtype of the isolates, respectively. Surveillance identified a total of 408 cases of meningococcal meningitis, with a case fatality rate of 8% (32/397). The mean annual incidence for the 304 culture-positive cases residing in metropolitan Salvador was 1.71 cases per 100,000 population. Infants <1 year old presented the highest incidence (14.7 cases per 100,000 population). Of the 377 serogrouped isolates, 82%, 16%, 2% and 0.3% were serogroups B, C, W135 and Y, respectively. A single serotype:serosubtype (4,7:P1.19,15) accounted for 64% of all cases. Continued surveillance is necessary to characterise strains and to define future prevention and control strategies.

Haemophilus influenzae meningitis 5 years after introduction of the Haemophilus influenzae type b conjugate vaccine in Brazil.

The long-term impact of Haemophilus influenzae type b (Hib) conjugate vaccine, introduced throughout Latin America in the late 1990s, has not been evaluated. Active surveillance for H. influenzae meningitis was performed from August 9, 1996 to August 8, 2004 in Metropolitan Salvador, Brazil. Five years after the introduction of Hib conjugate vaccine, Hib meningitis incidence decreased from 2.39 to 0.06 cases per 100,000 population (98%) overall, and from 60.9 to 3.1 cases per 100,000 population (95%) in children <1 year of age. A transient serotype replacement phenomenon was observed associated with a small increase of meningitis due to two H. influenzae type a clonal groups. These findings indicate that Hib immunization campaign has led to the virtual elimination of Hib disease in this region.

The cause of urinary symptoms among Human T Lymphotropic Virus Type I (HLTV-I) infected patients: a cross sectional study.

BACKGROUND: HTLV-I infected patients often complain of urinary symptomatology. Epidemiological studies have suggested that these individuals have a higher prevalence and incidence of urinary tract infection (UTI) than seronegative controls. However, the diagnosis of UTI in these studies relied only on patient information and did not require confirmation by urine culture. The purpose of this study was to investigate the role of urinary tract infection (UTI) as the cause of urinary symptoms in HTLV-I infected patients. METHODS: In this cross sectional study we interviewed, and cultured urine from, 157 HTLV-I seropositive individuals followed regularly at a specialized clinic. All patients were evaluated by a neurologist and classified according to the Expanded Disability Status Scale (EDSS). Urodynamic studies were performed at the discretion of the treating physician. RESULTS: Sixty-four patients complained of at least one active urinary symptom but UTI was confirmed by a positive urine culture in only 12 of these patients (19%); the majority of symptomatic patients (81%) had negative urine cultures. To investigate the mechanism behind the urinary complaints in symptomatic individuals with negative urine cultures, we reviewed the results of urodynamic studies performed in 21 of these patients. Most of them (90.5%) had abnormal findings. The predominant abnormalities were detrusor sphincter hyperreflexia and dyssynergia, findings consistent with HTLV-I-induced neurogenic bladder. On a multivariate logistic regression, an abnormal EDSS score was the strongest predictor of urinary symptomatology (OR 9.87, 95% CI 3.465 to 28.116, P < 0.0001). CONCLUSION: Urinary symptomatology suggestive of UTI is highly prevalent among HTLV-I seropositive individuals but true UTI is responsible for the minority of cases. We posit that the main cause of urinary symptoms in this population is neurogenic bladder. Our data imply that HLTV-I infected patients with urinary symptomatology should not be empirically treated for UTI but rather undergo urine culture; if a UTI is excluded, further investigation with urodynamic studies should be considered.

Antimony plus recombinant human granulocyte-macrophage colony-stimulating factor applied topically in low doses enhances healing of cutaneous Leishmaniasis ulcers: a randomized, double-blind, placebo-controlled study.

Cutaneous leishmaniasis (CL) requires 2-6 months to heal. In an effort to reduce this healing time, we studied topically applied granulocyte-macrophage colony-stimulating factor (GM-CSF) as an adjunct to antimonial therapy. Ten patients received antimony plus topical GM-CSF, and 10 patients received antimony plus placebo (saline). GM-CSF was diluted for topical use and was applied 3 times weekly for 3 weeks (1-2 microg/cm2/lesion). The mean +/- SD healing time was 43 +/- 14 days in the GM-CSF group and was 104+/-79 days in the placebo group (P=.043). Ten (100%) of 10 patients in the GM-CSF group healed within 60 days, compared with 5 (50%) of 10 patients in the placebo group. Two of the patients in the placebo group required retreatment with antimony. In conclusion, topically applied GM-CSF is effective in the management of CL.

Prevention of Haemophilus influenzae type b (Hib) meningitis and emergence of serotype replacement with type a strains after introduction of Hib immunization in Brazil.

Surveillance for Haemophilus influenzae meningitis cases was performed in Salvador, Brazil, before and after introduction of H. influenzae type b (Hib) immunization. The incidence of Hib meningitis decreased 69% during the 1-year period after initiation of Hib immunization (from 2.62 to 0.81 cases/100,000 person-years; P<.001). In contrast, the incidence for H. influenzae type a meningitis increased 8-fold (from 0.02 to 0.16 cases/100,000 person-years; P=.008). Pulsed-field gel electrophoretic analysis demonstrated that H. influenzae type a isolates belonged to 2 clonally related groups, both of which were found before Hib immunization commenced. Therefore, Hib immunization contributed to an increased risk for H. influenzae type a meningitis through selection of circulating H. influenzae type a clones. The risk attributable to serotype replacement is small in comparison to the large reduction in Hib meningitis due to immunization. However, these findings highlight the need to maintain surveillance as the use of conjugate vaccines expands worldwide.

Antimicrobial resistance in Haemophilus influenzae isolated during population-based surveillance for meningitis in Salvador, Brazil.

Antimicrobial susceptibility was determined for 150 Haemophilus influenzae isolates obtained during population-based surveillance for meningitis in Salvador, Brazil. Ten (6.7%) isolates were resistant to ampicillin and chloramphenicol. Of these, two isolates, a beta-lactamase and non-beta-lactamase producer, were resistant to amoxacillin-clavulinic acid. These findings indicate that present antibiotic regimens in Brazil may not be appropriate for the treatment of H. influenzae meningitis.

Population-based survey of antimicrobial susceptibility and serotype distribution of Streptococcus pneumoniae from meningitis patients in Salvador, Brazil.

Penicillin-nonsusceptible strains were isolated from 15% of 303 individuals with pneumococcal meningitis identified during a 4-year surveillance study in Salvador, Brazil. The estimated rate of coverage of the seven-valent conjugate vaccine was 74% among patients <5 years of age and 94% among those infected with nonsusceptible isolates, indicating that the use of conjugate vaccines may be an approach to the control of emerging penicillin resistance in Brazil.