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Rekha Pai

Publications and source records attributed to Rekha Pai.

8 recordsLinked to original sources
PubMed2006

Sequential multiplex PCR approach for determining capsular serotypes of Streptococcus pneumoniae isolates.

Accurate serotyping is essential to monitor the changes in the seroepidemiology of Streptococcus pneumoniae. We devised a simple and schematic sequence-based system of seven multiplex PCRs, in a sequence order based upon Active Bacterial Core surveillance (ABCs) serotype distribution during 2002 to 2003, to reliably deduce specific pneumococcal serotypes. A total of 421 isolates from ABCs were randomly chosen to evaluate this system. Two hundred twenty-nine of the isolates (54.3%) were specifically assigned 1 of 17 serotypes by the multiplex PCR system, with the results in complete concordance with conventional serotyping. One hundred seventy-two additional isolates (40.9%) were assigned to 11 specific sets of 2 to 4 serotypes that with one exception (serotypes 6A and 6B) consisted of the single frequently occurring targeted serotype and 1 to 3 additional rare serotypes primarily within the same serogroup as the targeted serotype. Only 20 isolates (4.8%) could not be assigned specific serotypes or serotype sets, since they were either of rare serotypes not included in the assay design or were nonserotypeable. Overall, we found this system to be highly reliable, with the potential to greatly reduce our reliance upon conventional serotyping. Especially important is the capability of this system to give serotype-determining potential to any facility that lacks the expensive typing sera and expertise needed for conventional serotyping yet has the modest equipment necessary for DNA amplification and electrophoresis.

Bacterial Capsules↗
PubMed2006

Pre- and postvaccination clonal compositions of invasive pneumococcal serotypes for isolates collected in the United States in 1999, 2001, and 2002.

Monitoring of serotypes and their clonal associations is critical as pneumococci adapt to the selective pressures exerted by the pneumococcal seven-valent conjugate vaccine (PCV7). We genotyped 1,476 invasive isolates from the Active Bacterial Core surveillance (705 [89.8%] of the isolates were obtained from children <5 years of age, and 771 [18.4%] of the isolates were obtained from individuals >5 years of age) in 2001 and 2002 (after the introduction of PCV7). The data were compared to the results for 1,168 invasive isolates (855 [83.9%] of the isolates were from children <5 years of age) collected in 1999. Among children <5 years of age, the incidence of invasive disease due to non-PCV7 serogroups together with serogroup 19A increased (P < 0.001). Eighty-three clonal sets, representing 177 multilocus sequence types (STs), were compiled from the 3-year isolate set. Among the non-PCV7 serogroups, newly emerging clones were uncommon; and a significant expansion of already established clones occurred for serotypes 3 (ST180), 7F (ST191), 15BCF (ST199), 19A (ST199), 22F (ST433), 33F (ST662), and 38 (ST393). However, additional minor clonal types within serotypes 1, 6A, 6B, 7C, 9N, 10A, 12F, 14, 15B/C, 17F, 19A, 19F, 20, 22F, and 33F that were absent in 1999 were found during 2001 and 2002. Although 23 clonal sets exhibited multiple serotypes, for most serotypes there were either no changes or modest changes in clonal compositions since the introduction of PCV7. The only example of an identical ST shared between non-PCV7 and PCV7 or PCV7-related serotypes was ST199; however, ST199 was prevalent within serotypes 15B/C and 19A before and after PCV7 introduction. Continued genotypic surveillance is warranted, since certain clones not targeted by PCV7 are expanding, and their emergence as significant pathogens could occur with maintained vaccine pressure.

Adult↗
PubMed2005

Postvaccine genetic structure of Streptococcus pneumoniae serotype 19A from children in the United States.

BACKGROUND: The introduction of the 7-valent conjugate pneumococcal vaccine (PCV7) in children may result in serotype replacement. We estimated the rate of increase of invasive pneumococcal disease (IPD) caused by serotype 19A in children <5 years old and determined the genetic composition of these isolates. METHODS: Cases of IPD between July 1999 and June 2004 were identified through the Active Bacterial Core Surveillance. Serotype 19A isolates obtained from children <5 years old between January 2003 and June 2004 were characterized by serotyping, antibiotic susceptibility testing, and pulsed-field gel electrophoresis (PFGE). Select isolates representing homologous PFGE clusters were subjected to multilocus sequence typing, and eBURST was used to delineate clonal groups. RESULTS: Between July 1999 and June 2004, the overall rate of IPD decreased from 23.3 to 13.1 cases/100,000 population (P<.00001). In children <5 years old, the rate decreased from 88.7 to 22.4 cases/100,000 population (P<.00001), whereas the rate in persons > or =5 years old decreased from 18.4 to 12.4 cases/100,000 population (P<.0001). The rate of serotype 19A IPD in children <5 years old increased significantly from 2.6 cases/100,000 population in 1999-2000 to 6.5 cases/100,000 population in 2003-2004; this was accompanied by significant increases in penicillin nonsusceptibility (P=.008) and multidrug resistance (P=.002) among serotype 19A isolates. As was observed during the pre-PCV7 era, clonal complex (CC) 199 predominated within serotype 19A, representing approximately 70% of invasive serotype 19A isolates from children <5 years old during 2003-2004. New serotype 19A genotypes were observed during 2003-2004, including 6 CCs that were not found among pneumococcal serotype 19A isolates during surveillance in 1999. CONCLUSION: Serotype 19A is, at present, the most important cause of IPD by replacement serotypes, and it is increasingly drug resistant. CC199 is the predominant CC among type 19A serotypes in children <5 years old. Our data suggest that some of the increase in rates of infection with serotype 19A may be due to serotype switching within certain vaccine type strains.

Anti-Bacterial Agents↗
PubMed2005

Clonal association between Streptococcus pneumoniae serotype 23A, circulating within the United States, and an internationally dispersed clone of serotype 23F.

Streptococcus pneumoniae is an important pathogen in the United States and is associated with significant morbidity and mortality. Since the introduction of the seven-valent conjugate vaccine, a significant decline in pneumococcal disease has been reported. However, surveillance for pneumococcal disease remains essential, as the extent of cross protection against vaccine-related serotypes is still unclear. Further, any increase in non-vaccine-related serotypes also needs monitoring. We report on a new clonal association between a vaccine-related serotype, serotype 23A, obtained as part of the Active Bacterial Core surveillance, with an established internationally dispersed Pneumococcal Molecular Epidemiology Network (PMEN) clone, clone Colombia(23F)-26. Sixty-two isolates of serotype 23A collected from sterile sites during a 2-year period (2002 and 2003) were characterized. Twenty-one (34%) isolates were penicillin nonsusceptible, although none were fully resistant. Pulsed-field gel electrophoresis and multilocus sequence typing analysis showed that 24 (39%) of the serotype 23A isolates shared either genetic identity or high genetic relatedness with PMEN clone Colombia(23F)-26. Extensive variability was noted within the sequenced region of pbp2b in two penicillin-nonsusceptible isolates as well as in PMEN clone Colombia(23F)-26, suggesting that these isolates probably acquired penicillin resistance independently. The emergence of such new serotype and genotype associations highlights the dynamic nature of the pneumococcal population, necessitating continuous monitoring in the post-vaccine era.

Adolescent↗
PubMed2005

Use of pyrosequencing to differentiate Streptococcus pneumoniae serotypes 6A and 6B.

Accurate serotyping of Streptococcus pneumoniae remains important to monitor the changes in seroepidemiology of the organism over time. Though several PCR-based systems have been developed for this purpose, the cross-reactivity within serogroups often limits discrimination between types. All serogroup 6 isolates can be identified using a multiplex PCR system; however, due to the high sequence homology between the cps-6B and cps-6A loci, serotypes 6A and 6B cannot be differentiated by this method. We describe the use of pyrosequencing to reliably differentiate between serotypes 6A and 6B using a previously described single nucleotide polymorphism at codon 195 of the cps locus wciP gene. We observed complete concordance between capsular serotyping results and wciP pyrosequencing among 210 isolates examined, indicating that pyrosequencing is a rapid and accurate technique for deducing serotypes 6A and 6B.

Bacterial Capsules↗
PubMed2003

Comparison of disc diffusion & E test methods with agar dilution for antimicrobial susceptibility testing of Haemophilus influenzae.

BACKGROUND & OBJECTIVES: Reliable methods of detection of antimicrobial resistance are of paramount importance in the treatment and management of infections caused by Haemophilus influenzae. The objective of the present study was to compare and evaluate the performance of disc diffusion and E test (Epsilometer test) with agar dilution method for antimicrobial susceptibility testing of H. influenzae. METHODS: A total of 46 isolates of H. influenzae from various invasive sites were included as test strains. Antimicrobial susceptibility testing was performed by Kirby-Bauer disc diffusion method for ampicillin, chloramphenicol, trimethoprim-sulphamethoxazole (TMP-SMZ) and cefotaxime. Minimum Inhibitory Concentration (MIC) determination was performed by E test and agar dilution for the same set of antimicrobials. All tests were performed on Haemophilus test medium (HTM). RESULTS: Disc diffusion showed a very major (2%) and minor (4%) interpretative error with TMP-SMZ and minor interpretative errors to ampicillin (13%) and chloramphenicol (24%) when compared to agar dilution method. E test produced only minor interpretative errors to chloramphenicol (7%) and TMP-SMZ (2%) and no interpretative errors with ampicillin and cefotaxime as against agar dilution. E test showed good agreement with agar dilution for each of the antimicrobial tested. INTERPRETATION & CONCLUSION: Disc diffusion test may be used as a preliminary screen for susceptibility testing of H. influenzae. E test is simple, easy to perform and a reliable method for determination of resistance in H. influenzae. However its cost and limited availability in India may limit its use. The reference agar dilution method can be used reliably in routine susceptibility testing of H. influenzae.

Agar↗
PubMed2002

Fully integrated on-chip electrochemical detection for capillary electrophoresis in a microfabricated device.

Microfabricated lab-on-a-chip devices employing a fully integrated electrochemical (EC) detection system have been developed and evaluated. Both capillary electrophoresis (CE) channels and all CE/EC electrodes were incorporated directly onto glass substrates via traditional microfabrication techniques, including photolithographic patterning, wet chemical etching, DC sputtering, and thermal wafer bonding. Unlike analogous CE/EC devices previously reported, no external electrodes were required, and critical electrode characteristics, including size, shape, and placement on the microchip, were established absolutely by the photolithography process. For the model analytes dopamine and catechol, detection limits in the 4-5 microM range (approximately 200 amol injected) were obtained with the Pt EC electrodes employed here, and devices gave stable analytical performance over months of usage.

Catechols↗