Rosemeire Cobo Zanella

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.

Decreased Point Prevalence of Haemophilus influenzae Type b (Hib) Oropharyngeal Colonization by Mass Immunization of Brazilian Children Less Than 5 Years Old with Hib Polyribosylribitol Phosphate Polysaccharide—Tetanus Toxoid Conjugate Vaccine in Combination with Diphtheria-Tetanus Toxoids—Pertussis Vaccine

A protective herd effect has been described after susceptible populations of children are vaccinated with conjugate Haemophilus influenzae type b (Hib). Hib carriage was studied in children aged 6-24 months attending day care centers in two cities in southern Brazil (Curitiba and Porto Alegre). In Curitiba, routine immunization with Hib polyribosylribitol phosphate polysaccharide-tetanus toxoid conjugate vaccine (PRP-T) in combination with diphtheria-tetanus toxoids-pertussis vaccine (PRP-T/DTP) has been offered since September 1996; DTP vaccine alone is routinely given in Porto Alegre. Children in Porto Alegre (n=643) were 8 times less likely to have received adequate Hib vaccination and 4 times more likely to be Hib carriers than children in Curitiba (n=647; i.e., point prevalence of oropharyngeal colonization, 4.8% vs. 1.2%). Point prevalence of carriage with non-type b or other nontypeable Hi was similar in children of both cities. There was a vaccination effect on carriage rates in children who received a primary 3-dose series, independent of the booster dose, suggesting that a booster may be unnecessary to induce population protection.

Multilocus Variable-Number Tandem-Repeat Polymorphism among Brazilian Enterococcus faecalis Strains

ABSTRACT Multilocus variable-number tandem-repeat analysis (MLVA) for seven genomic loci was developed for Enterococcus faecalis. MLVA and pulsed-field gel electrophoresis (PFGE) resulted in 37 and 31 genotypes among 83 strains, respectively. Both typing schemes were highly concordant (90.4%). MLVA is an excellent alternative to PFGE.

Phenotypic and genotypic characterization of VanA Enterococcus isolated during the first nosocomial outbreak in Brazil.

We report the phenotypic and genotypic characterization of 50 VanA Enterococcus clinical isolates from infected patients and 97 isolates from colonized patients obtained during a nosocomial outbreak in a single hospital in São Paulo, Brazil during 1998. The identification of strains to the species level by conventional biochemical and phenotypic tests and by multiplex PCR assay had 100% agreement. Both E. faecalis and E. faecium were isolated from patients during this outbreak. The vanA genotype was confirmed by PCR. Antibiotic susceptibility testing showed that E. faecium isolates are generally less susceptible to antibiotics than E. faecalis. By PCR, 24 of 26 VRE strains tested carried the Tn1546 element. Pulsed-field gel electrophoresis identified five distinct patterns for E. faecalis (A, B, C, D, E) and three for E. faecium (M, N, and O). A single PFGE pattern was identified in the majority of strains of each species and does not discriminate between case and carrier isolates.

Evaluation of Methodology for Serotyping Invasive and Nasopharyngeal Isolates of Haemophilus influenzae in the Ongoing Surveillance in Brazil

ABSTRACT To assess the magnitude of discrepant results obtained by routine Haemophilus influenzae serotyping, 258 isolates, collected by the epidemiological surveillance system in Brazil from individuals with invasive diseases or carriage, were evaluated by two slide agglutination (SlAg) methods: SlAg method 1, by which strains were initially screened with a serotype b-specific antiserum, and SlAg method 2, by which strains were tested against all serotype-specific antisera in parallel. Investigators comparing results of the two SlAg methods with those obtained by capsule type-specific PCR were blinded to the method used. The serotype prevalence rates found by the three methods were significantly different, involving discrepancies mainly between serotype b and noncapsulated (NC) isolates. For invasive isolates (n = 131), the overall agreement rate between SlAg method 1 or 2 and PCR was 68.0 or 88.3%, respectively, whereas for colonizing isolates (n = 127) the corresponding rate was 46.5 or 94.2%, respectively. SlAg method 2 improved the ascertainment of serotypes over that obtained with SlAg method 1, demonstrating good correlation with PCR. Use of the polyvalent antiserum as a screening reagent for SlAg for invasive and colonizing isolates showed poor discriminatory power, with a sensitivity of 65.8% and a specificity of 91.7%. We stress the importance of using a well-standardized SlAg methodology and suggest that reference laboratories should utilize PCR routinely to confirm SlAg results and to check all nonspecific SlAg reactions and apparent NC isolates by SlAg in order to provide reliable data on the prevalence of H. influenzae serotypes in the H. influenzae type b vaccine era.

Effect of 10-valent pneumococcal conjugate vaccine on nasopharyngeal carriage of Streptococcus pneumoniae and Haemophilus influenzae among children in São Paulo, Brazil

In March 2010, Brazil introduced the 10-valent pneumococcal conjugate vaccine (PCV10) in the routine infant immunization program using a 4-dose schedule and catch-up for children <23months. We investigated PCV10 effect on nasopharyngeal carriage with vaccine-type Streptococcus pneumoniae (Spn) and non-typeable Haemophilus influenzae (NTHi) among children in São Paulo city. Cross-sectional surveys were conducted in 2010 (baseline) and 2013 (post-PCV10). Healthy PCV-naïve children aged 12-23months were recruited from primary health centers during immunization campaigns. Nasopharyngeal swabs were collected and tested for Hi; for Spn, all baseline and a stratified random sample of 400 post-PCV10 swabs were tested. We compared vaccine-type Spn and NTHi carriage prevalence pre-/post-PCV10, and used logistic regression to estimate PCV10 effectiveness (1-adjusted odds ratio×100%). Overall 501 children were included in the baseline and 1167 in the post-PCV10 survey (including 400 tested for Spn). Spn was detected in 40.3% of children at baseline and 48.8% post-PCV10; PCV10 serotypes were found in 19.8% and 1.8% respectively, representing a decline of 90.9% (p<0.0001). Carriage of vaccine-related serotypes increased (10.8-21.0%, p<0.0001), driven primarily by a rise in serotype 6C (1.8-11.2%, p<0.0001); carriage of serotypes 6A and 19A did not significantly change. PCV10 effectiveness (4 doses) against vaccine-type carriage was 97.3% (95% confidence interval 88.7-99.3). NTHi prevalence increased from 26.0% (130/501) to 43.6% (509/1167, p<0.0001); PCV10 vaccination seemed significantly associated with NTHi carriage, even after adjusting for other known risk factors. Carriage with PCV10 serotypes among toddlers declined dramatically following PCV10 introduction in São Paulo, Brazil. No protection of PCV10 against NTHi was observed. Our findings contribute to a growing body of evidence of PCV10 impact on vaccine-type carriage and highlight the importance of PCV10 as a tool to reduce the burden of pneumococcal disease in Brazil and globally.

Changes in serotype distribution of Haemophilus influenzae meningitis isolates identified through laboratory-based surveillance following routine childhood vaccination against H. influenzae type b in Brazil.

Following routine childhood vaccination against Haemophilus influenzae type b (Hib) disease in Brazil in 1999, passive laboratory surveillance reported increasing numbers of non-b serotypes and nontypeable H. influenzae (NTHi) from meningitis cases. To characterize this increase, we analyzed data on 3910 H. influenzae isolated from cerebrospinal fluid or blood from meningitis cases that were sent to the national reference laboratory for serotyping from 1990 to 2008. Hib accounted for 98% of H. influenzae meningitis isolates received during 1990-1999 versus 59% during 2000-2008, while non-b serotypes increased from 1% to 19% and NTHi increased from 2% to 22% of H. influenzae isolates received during the two periods. Higher proportions of non-b serotypes and NTHi than Hib were isolated from blood rather than cerebrospinal fluid. Estimated incidence rates for H. influenzae meningitis for Sao Paulo state remained below 1 case per million population during 2000-2008, although annual incidence of NTHi meningitis (mean, 0.03 cases per 100,000 population) increased in several age groups. Changes in surveillance for H. influenzae following introduction of Hib conjugate vaccine likely contributed to increased numbers of non-b and nontypeable H. influenzae meningitis isolates received at the national reference laboratory.

Molecular assessment of invasive Streptococcus pneumoniae serotype 1 in Brazil: evidence of clonal replacement

In Brazil, serotype 1 Streptococcus pneumoniae is one of the most prevalent causes of severe infection. This study investigated the genetic relatedness of 134 serotype 1 isolates obtained from invasive diseases during the period 1977-2005. Molecular typing by PFGE revealed two major lineages using visual inspection and computer analysis. Type A comprised 94 isolates (70.2 %) with four subtypes, whereas type B comprised 40 isolates (29.8 %) with eight subtypes. Subtype A3, the most frequent genotype, accounting for 65 % of the total isolates, was identified as a representative of clone Sweden(1)-40 (ST304). Type B was predominant in the period 1977-1988. In contrast, an increase in the type A lineage was detected from 1990 in Brazil, significantly associated with isolates recovered from pneumonia cases and from young patients. This study clearly established a temporal switch between two lineages of S. pneumoniae serotype 1 in Brazil, with a wide dispersion of clone Sweden(1)-40 in recent years.

Characterization of Haemophilus influenzae Isolated from Invasive Disease in Brazil from 1990 to 1999

The Haemophilus influenzae serotype b (Hib) conjugate vaccine was introduced in the National Immunization Program in Brazil in the second half of 1999. A retrospective analysis on serotypes, biotypes, and antimicrobial resistance of Hi invasive strains obtained through Hi survey was conducted to document the characteristics of this pathogenic agent during a decade prior the use of Hib vaccine. A total 3,204 strains from 1990 to 1999 were studied, being 88.2% isolated from cerebrospinal fluid, 10.7% from blood, and 1.1% from pleural fluid. The rate of 90.9% of strains was obtained from children up to 4 years old, and the age group >6 months old to 1 year was the higher risk to Hi infection. Type b was, by far, the most common type (97.8%), followed in frequency by type a (0.5%); only 1.5% was a nontypable strain. Biotypes I and II accounted for 97.8% of isolates. Resistance to ampicillin (AM) and chloramphenicol (CO) was detected at rates of 18.1% and 19.1%, respectively, whereas simultaneous resistance to AM and CO was identified in 13.9% of strains. Total concordance was found between AM resistance and beta-lactamase production. No strain showed resistance to ceftriaxone and rifampicin. In conclusion, the data generated through this laboratory-based surveillance should serve as a reference for assessing the impact of Hib vaccination and to detect changes on the pattern of Hi diseases in the country.

Carriage of Haemophilus influenzae among Brazilian children attending day care centers in the era of widespread Hib vaccination

Haemophilus influenzae type b vaccine was introduced into the Immunization Program of Brazil in 1999 and no study has evaluated the impact of Hib vaccination in H. influenzae carriage so far. In June 2010, Brazil introduced the 10-valent pneumococcal nontypeable H. influenzae (NTHi) conjugate vaccine (PHiD-CV). We investigated the prevalence of encapsulated H. influenzae and NTHi isolates in nasopharyngeal samples of 1192 children attending day-care centers in Goiânia, central Brazil. H. influenzae carriage rate was 32.1% and 38.4% of them carried β-lactamase TEM-1 gene. Serotype f (4.6%) was the most frequent encapsulated isolate, type b was recovered in only 0.7% and carriage rate of NTHi was 23.3%. Recurrent acute otitis media and NTHi were independently associated with colonization by β-lactamase producing H. influenzae. Changes in frequency of H. influenzae carriage isolates should be carefully monitored to assess the impact of the PHiD-CV on NTHi carriage in young children.