Ninety-five colonizing isolates and 74 invasive isolates of Streptococcus agalactiae from Kenyan adults were characterized by using capsular serotyping and multilocus sequence typing. Twenty-two sequence types clustering into five clonal complexes were found. Data support the view that S. agalactiae isolates belonging to a limited number of clonal complexes are invasive in adults worldwide

Bisharat N.

Blancas D.

Edwards M. S.

Farley M. M.

Imperi M.

Jolley K. A.

Lefevre J. C.

Phares C. R.

Schwartz B.

Sendi P.

Skoff T. H.

Sørensen U. B. S.

Valkenburg-van den Berg A. W.

Verani J. R.

PubMed

ABSTRACT
          
            Ninety-five colonizing isolates and 74 invasive isolates of
            Streptococcus agalactiae
            from Kenyan adults were characterized by using capsular serotyping and multilocus sequence typing. Twenty-two sequence types clustering into five clonal complexes were found. Data support the view that
            S. agalactiae
            isolates belonging to a limited number of clonal complexes are invasive in adults worldwide.
          </jats:p

Charlotte A. Huber

Francis McOdimba

Valentin Pflueger

Claudia A. Daubenberger

Gunturu Revathi

Crossref

Journal of Clinical Microbiology

Characterization of Invasive and Colonizing Isolates of Streptococcus agalactiae in East African Adults

Huber, Charlotte A.

McOdimba, Francis

Pflueger, Valentin

Daubenberger, Claudia A.

Revathi, Gunturu

University of Queensland eSpace

JOURNAL OF CLINICAL MICROBIOLOGY, Oct. 2011, p. 3652–3655 Vol. 49, No. 10
0095-1137/11/$12.00 doi:10.1128/JCM.01288-11
Copyright © 2011, American Society for Microbiology. All Rights Reserved.
Characterization of Invasive and Colonizing Isolates of
Streptococcus agalactiae in East African Adults
Charlotte A. Huber,1 Francis McOdimba,3 Valentin Pflueger,2
Claudia A. Daubenberger,1‡ and Gunturu Revathi3‡*
Swiss Tropical and Public Health Institute and University of Basel, Department of Medical Parasitology and Infection Biology,
Socinstrasse 57, 4002 Basel, Switzerland1; Mabritec SA, Lo¨rracherstrasse 50, P.O. Box 320, 4125 Riehen, Switzerland2; and
Aga Khan University Hospital, Nairobi, 3rd Parklands Avenue, P.O. Box 30270-00100, Nairobi, Kenya3
Received 28 June 2011/Returned for modification 28 July 2011/Accepted 12 August 2011
Ninety-five colonizing isolates and 74 invasive isolates of Streptococcus agalactiae from Kenyan adults were
characterized by using capsular serotyping and multilocus sequence typing. Twenty-two sequence types clus-
tering into five clonal complexes were found. Data support the view that S. agalactiae isolates belonging to a
limited number of clonal complexes are invasive in adults worldwide.
Streptococcus agalactiae, or group B streptococcus (GBS),
regarded mainly as a pathogen of neonates and pregnant
women (15), is increasingly affecting nonpregnant adults (19).
In adults, S. agalactiae causes skin and soft tissue infections,
bacteremia, urinary tract infections, pneumonia, osteomyelitis,
meningitis, endocarditis, and streptococcal toxic shock syn-
drome (2, 6, 18, 20). Risk factors associated with invasive GBS
in adults are old age, diabetes mellitus, neurologic diseases,
cirrhosis or other liver diseases, stroke, breast cancer, and
renal failure (7, 22, 23). S. agalactiae colonizes the lower gas-
trointestinal and genitourinary tracts of 30 to 50% of healthy
adults (26), and an estimated 20 to 30% of all pregnant women
are carriers (25). S. agalactiae can be isolated from vaginal or
rectal swabs, and prenatal screening for colonization of preg-
nant women is recommended (27). The objective of this study
was to analyze the epidemiology of GBS in East African adults
by using multilocus sequence typing (MLST) (13) and capsular
serotyping (10), with a focus on investigating possible correla-
tions between clonal complexes of GBS in invasive or coloniz-
ing isolates.
The Aga Khan University Hospital in Nairobi (AKUH,N) is
a tertiary care, referral university hospital in Nairobi, Kenya.
GBS were cultivated at the Division of Microbiology of
AKUH,N from specimens derived from both inpatients and
outpatients between January 2007 and June 2010. Swabs from
outreach collection points were transported in Stuart’s trans-
port medium to the hospital. Blood cultures were routinely
performed using Plus-aerobic/F, Plus-anaerobic/F, and Peds
Plus Bactec 9120 media (Becton Dickinson) in combination
with commercially available biphasic culture medium bottles
(Roche Diagnostics, France). Single GBS colonies were picked
from 5% sheep blood agar plates (Oxoid, United Kingdom),
and identification of GBS was performed using colony mor-
phology, Gram staining, CAMP test, the API Streptococcus
identification kit (Bio-Merieux, France), and the latex aggluti-
nation test using a bacterial meningitis kit with specific group
B latex (Wellcogen kit from Remel Europe Ltd.). Antimicro-
bial susceptibility testing was performed, and results were in-
terpreted according to the guidelines of the Clinical and
Laboratory Standards Institute (CLSI) (5). Streptococcus pneu-
moniae ATCC 49619 was used for quality control. All GBS
isolates were found to be susceptible to penicillin and cepha-
losporin. GBS isolates analyzed here were selected according
to availability of stocked isolates and accessibility of patient
files. One isolate per patient was included in this study. Clinical
information was obtained by review of patient files. Ethical
clearance for this study was obtained from the Ethikkommis-
sion beider Basel (EKBB) and the AKUH scientific and ethical
committee.
For MLST, GBS isolates were grown overnight at 37°C on
plates with Columbia agar enriched with 5% sheep blood
(Oxoid). Genomic DNA extraction, MLST PCR amplification,
and sequencing of seven housekeeping genes were performed
as described previously (13). Novel alleles of the housekeeping
genes were amplified and sequenced twice and submitted to
Nicola Jones, Oxford, United Kingdom, for allele assignment.
The sequence types (STs) were determined by the S. agalactiae
MLST website (http://pubmlst.org/sagalactiae/) (12). The neigh-
bor-joining tree was generated from the allelic profiles ob-
tained using Start2 (11). The multiplex PCR assay for identi-
fication of capsular serotypes was performed as described
previously (10). To assess if any clonal complex is associated
with colonizing or invasive isolates, the distribution of each
clonal complex was examined against random distribution by
using the Pearson chi-square test. Association between
clonal complex and capsular serotype distributions was as-
sessed by using Fisher’s exact test. Statistical analyses were
performed using STATA version 10 (Stata Corporation,
College Station, TX).
Ninety-five colonizing GBS isolates were derived from vag-
inal swabs collected during routine prenatal screening of preg-
nant women. Out of the 74 invasive isolates, 40 (54%) were
derived from urine, 10 (13.5%) from blood, 8 (10.8%) from
pus, and 16 (21.6%) from other body locations. Thirty-seven
(50%) of the GBS from urine samples originated from women.
* Corresponding author. Mailing address: Aga Khan University
Hospital, Nairobi, 3rd Parklands Avenue, Nairobi, Kenya. Phone: 254-
20-3662226. Fax: 254-20-3741749. E-mail: gunturu.revathi@aku.edu.
‡ These authors contributed equally.
 Published ahead of print on 24 August 2011.
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Twenty-one of these nonpregnant female patients had re-
peated attacks of symptomatic urinary tract infection with sig-
nificant pyuria and heavy vaginal colonization. Nine patients
were antenatal symptomatic women with significant pyuria and
bacteriuria. Two male patients with chronic GBS urinary tract
infection suffered from prostatic hypertrophy. One elderly
male patient suffered from catheter-associated GBS urinary
tract infection due to neurogenic bladder. This patient devel-
oped Pseudomonas aeruginosa bacteriuria after GBS subsided
under antibiotic treatment (8, 21). Five of the patients with
positive blood cultures were elderly adults with comorbidities,
including diabetes, renal failure, cardiac asthma, and malig-
nancies with primary GBS bacteremia. One adult male devel-
oped GBS bacteremia secondary to peritonitis and later pro-
gressed to renal failure subsequent to toxic shock. Out of the 8
pus isolates, 4 came from cases of osteomyelitis and one each
came from a diabetic foot, liver abscess, pelvic abscess aspirate,
and joint fluid. A fat tissue isolate was cultured from postmor-
tem samples from an elderly woman. Five isolates from ure-
thral swabs came from patients with nongonococcal urethritis
(16).
In total, 22 STs could be distinguished in this collection, with
the colonizing and invasive GBS isolates grouped into 16 STs
and 19 STs, respectively (Table 1). Isolates were grouped into
clonal complexes (CCs) by eBURST, relaxing the group defi-
nition to six out of seven shared alleles. Thirteen STs were
clustered in five CCs, namely, CC23, CC19, CC17, CC10, and
CC1 (Fig. 1). The most prevalent CC was CC23 (27.2%),
followed by CC17 (21.3%), CC10 (16.6%), CC19 (13.6%), and
CC1 (12.4%). Eighty-eight percent of the isolates could be
grouped into one of these five CCs. These five CCs are also
highly prevalent in other GBS collections representing either
worldwide collections (24) or investigations performed in Swe-
den (17), Italy (9), England (14), the United States (3), the
Central African Republic, and Senegal (4) as well as Israel (1).
ST-26, which represented 15% of all identified GBS isolates in
the Central African Republic and Senegal (4), was found three
times in our collection only. Singletons identified were ST-3,
ST-4, ST-24, ST-26 ST-103, ST-327, ST-328, ST-485, and ST-
486 (Table 1). Five novel STs named ST-484, ST-485, ST-486,
ST-492, and ST-501 were detected. ST-485 is a double-locus
variant and ST-486 is a single-locus variant of ST-103. ST-484
is a single-locus variant of ST-17, while ST-492 and ST-501 are
single-locus variants of ST-23. All novel STs were found at
least once in clinical isolates, and ST-484 and ST-486 were
additionally found in colonizing isolates. The most prevalent
novel ST was ST-484, found in 6 case isolates and 2 colonizing
isolates. ST-4, ST-103, ST-327, ST-485, ST-492, and ST-501
were found exclusively in invasive GBS isolates. ST-3, ST-24,
and ST-8 could be detected only among the colonizing GBS
TABLE 1. Overview of the GBS sequence types and capsular serotypes obtained in the current study
CC and/or ST
No. of isolates Capsular serotype(s) (no. of isolates)
Total Colonizing Invasive Total Colonizing Invasive
CC1a
ST-1 16 9 7 V (15), Ia (1) V (9) V (6), Ia (1)
ST-2 5 4 1 II (4), V (1) II (4) V (1)
CC10a
ST-8 5 5 0 Ib (5) Ib (5)
ST-10 20 10 10 Ib (18), II (2) Ib (9), II (1) Ib (9), II (1)
ST-12 3 1 2 V (2), Ib (1) V (1) V (1), Ib (1)
CC17a
ST-17 28 20 8 III (28) III (20) III (8)
ST-484 8 2 6 III (8) III (2) III (6)
CC19a
ST-19 8 6 2 III (5), V (3) III (5), V (1) V (2)
ST-28 6 3 3 II (6) II (3) II (3)
ST-182 9 4 5 III (9) III (4) III (5)
CC23a
ST-23 44 24 20 Ia (42), III (2) Ia (24) Ia (18), III (2)
ST-492 1 0 1 Ia (1) Ia (1)
ST-501 1 0 1 Ia (1) Ia (1)
Singletons
ST-3 1 1 0 IV (1) IV (1)
ST-4 1 0 1 Ia (1) Ia (1)
ST-24 1 1 0 Ia (1) Ia (1)
ST-26 3 2 1 V (3) V (2) V (1)
ST-103 1 0 1 Ia (1) Ia (1)
ST-327 1 0 1 V (1) V (1)
ST-328 4 2 2 V (4) V (2) V (2)
ST-485 1 0 1 Ia (1) Ia (1)
ST-486 2 1 1 Ia (2) Ia (1) Ia (1)
a Fisher’s exact test P value, 0.000 (comparison among CCs).
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bacteria (Table 1). Interestingly, out of the 10 isolates obtained
from blood, the novel ST-484, ST-485, ST-486, and ST-492
represented 30% (Table 2), supporting their high invasive po-
tential. The possible relationship between invasive or coloniz-
ing GBS and the five CCs was analyzed (Table 2). No statisti-
cally significant correlation between any of the five CCs and
invasive or colonizing GBS isolates could be described by using
Pearson’s chi-square test (P  0.926). However, here we com-
pare colonizing isolates from pregnant woman with invasive
isolates from nonpregnant adults.
The capsular serotypes detected here included Ia, Ib, II, III,
IV, and V. The most common serotype was serotype III
(30.8%), followed by serotype Ia (30.2%), serotype V (17.2%),
serotype Ib (14.2%), and serotype II (7.1%); one isolate was of
serotype IV (0.6%). Out of the 44 isolates identified as ST-23,
42 were serotype Ia. All ST-17 isolates carried serotype III, 18
FIG. 1. Neighbor-joining tree demonstrating the relationship between the 22 STs of the GBS identified in the current study. The
phylogenetic tree is based on the alleles of the seven housekeeping alleles sequenced and was constructed using the Start2 program
(http://pubmlst.org/software/analysis/start2/). The scale bar indicates distance as calculated by the number of mismatched loci.
TABLE 2. Sequence types and capsular serotypes of the GBS clinical isolates according to location of origin
CC and/or ST
No. of isolates Capsular serotype(s) (no. of isolates)
Total Urine Blood Pus Other Total Urine Blood Pus Other
CC1a
ST-1 7 4 1 2 V (6), Ia (1) V (4) Ia (1) V (2)
ST-2 1 1 V (1) V (1)
CC10a
ST-10 10 5 1 4 Ib (9), II (1) Ib (5) II (1) Ib (4)
ST-12 2 1 1 V (1), Ib (1) V (1) Ib (1)
CC17a
ST-17 8 7 1 III (8) III (7) III (1)
ST-484 6 3 2 1 III (6) III (3) III (2) III (1)
CC19a
ST-19 2 2 V (2) V (2)
ST-28 3 3 II (3) II (3)
ST-182 5 2 2 1 III (5) III (2) III (2) III (1)
CC23a
ST-23 20 10 2 3 5 Ia (18), III (2) Ia (9), III (1) Ia (2) Ia (2), III (1) Ia (5)
ST-492 1 1 Ia (1) Ia (1)
ST-501 1 1 Ia (1) Ia (1)
Singletons
ST-4 1 1 Ia (1) Ia (1)
ST-26 1 1 V (1) V (1)
ST-103 1 1 Ia (1) Ia (1)
ST-327 1 1 V (1) V (1)
ST-328 2 2 V (2) V (2)
ST-485 1 1 Ia (1) Ia (1)
ST-486 1 1 Ia (1) Ia (1)
a Pearson’s chi-square test P value, 0.926 (comparison among CCs).
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out of 20 ST-10 isolates had serotype Ib, 15 out of 16 ST-1
isolates expressed V, and all ST-484 and ST-182 isolates had
serotype III. Hence, a tight relationship between distinct cap-
sular serotypes and the CC could be observed (Fisher’s exact
test P  0.000) (Table 1). Of note, invasive GBS typed as
ST-23, ST-12, and ST-1 showed a higher variability in their
capsular serotypes than colonizing isolates (Table 1). In sum-
mary, our data show that the population structure of GBS
isolates in East African adults is similar to the GBS population
structure reported in industrialized countries. A number of
novel STs that share alleles with housekeeping genes of wide-
spread known STs were detected.
We are thankful to the team in the Division of Microbiology, De-
partment of Pathology, at the AKUH,N. The excellent assistance ren-
dered by Patricia Shivachi with data management is gratefully ac-
knowledged. The contributions made by Patricia Muthaura, Timona
Obura, Alfred Murage, Maria Carvaloh, K. K. Bal, Elizabeth Njoroge,
Jyothi Saha, and P. S. Patel are gratefully acknowledged. We are
thankful to Penelope Vounatsou for statistical support.
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Characterization of invasive and colonizing isolates of Streptococcus agalactiae in East African adults

95 colonizing and 74 invasive Streptococcus agalactiae isolates from Kenyan adults were characterized using capsular serotyping and multilocus sequence typing. 22 sequence types clustering into five clonal complexes were found. Data support the view that S. agalactiae isolates belonging to limited number of clonal complexes are invasive in adults worldwid

Huber, C. A.

McOdimba, F.

Pflueger, V.

Daubenberger, C. A.

Revathi, G.

edoc

Characterisation of invasive and colonizing isolates of Streptococcus agalactiae in East African adults

Huber, Charlotte

Daubenberger, Claudia

eCommons@AKU

eCommons@AKUPathology, East Africa Medical College, East AfricaOctober 2011Characterization of invasive and colonizing isolatesof Streptococcus agalactiae in East African adultsCharlotte HuberSwiss Tropical and Public Health Institute and University of BaselFrancis McOdimbaAga Khan UniversityValentin PfluegerMabritec SAClaudia DaubenbergerSwiss Tropical and Public Health Institute and University of BaselGunturu RevathiAga Khan UniversityFollow this and additional works at: http://ecommons.aku.edu/eastafrica_fhs_mc_patholPart of the Pathology CommonsRecommended CitationHuber, C., McOdimba, F., Pflueger, V., Daubenberger, C., Revathi, G. (2011). Characterization of invasive and colonizing isolates ofStreptococcus agalactiae in East African adults. Journal of Clinical Microbiology, 49(10), 3652-3655.Available at: http://ecommons.aku.edu/eastafrica_fhs_mc_pathol/3  Published Ahead of Print 24 August 2011. 10.1128/JCM.01288-11. 2011, 49(10):3652. DOI:J. Clin. Microbiol. Claudia A. Daubenberger and Gunturu RevathiCharlotte A. Huber, Francis McOdimba, Valentin Pflueger, African AdultsEastIsolates of Streptococcus agalactiae in Characterization of Invasive and Colonizinghttp://jcm.asm.org/content/49/10/3652Updated information and services can be found at: These include:REFERENCEShttp://jcm.asm.org/content/49/10/3652#ref-list-1at: This article cites 26 articles, 12 of which can be accessed freeCONTENT ALERTS more»articles cite this article), Receive: RSS Feeds, eTOCs, free email alerts (when newhttp://journals.asm.org/site/misc/reprints.xhtmlInformation about commercial reprint orders: http://journals.asm.org/site/subscriptions/To subscribe to to another ASM Journal go to:  on November 9, 2012 by guesthttp://jcm.asm.org/Downloaded from JOURNAL OF CLINICAL MICROBIOLOGY, Oct. 2011, p. 3652–3655 Vol. 49, No. 100095-1137/11/$12.00 doi:10.1128/JCM.01288-11Copyright © 2011, American Society for Microbiology. All Rights Reserved.Characterization of Invasive and Colonizing Isolates ofStreptococcus agalactiae in East African AdultsCharlotte A. Huber,1 Francis McOdimba,3 Valentin Pflueger,2Claudia A. Daubenberger,1‡ and Gunturu Revathi3‡*Swiss Tropical and Public Health Institute and University of Basel, Department of Medical Parasitology and Infection Biology,Socinstrasse 57, 4002 Basel, Switzerland1; Mabritec SA, Lo¨rracherstrasse 50, P.O. Box 320, 4125 Riehen, Switzerland2; andAga Khan University Hospital, Nairobi, 3rd Parklands Avenue, P.O. Box 30270-00100, Nairobi, Kenya3Received 28 June 2011/Returned for modification 28 July 2011/Accepted 12 August 2011Ninety-five colonizing isolates and 74 invasive isolates of Streptococcus agalactiae from Kenyan adults werecharacterized by using capsular serotyping and multilocus sequence typing. Twenty-two sequence types clus-tering into five clonal complexes were found. Data support the view that S. agalactiae isolates belonging to alimited number of clonal complexes are invasive in adults worldwide.Streptococcus agalactiae, or group B streptococcus (GBS),regarded mainly as a pathogen of neonates and pregnantwomen (15), is increasingly affecting nonpregnant adults (19).In adults, S. agalactiae causes skin and soft tissue infections,bacteremia, urinary tract infections, pneumonia, osteomyelitis,meningitis, endocarditis, and streptococcal toxic shock syn-drome (2, 6, 18, 20). Risk factors associated with invasive GBSin adults are old age, diabetes mellitus, neurologic diseases,cirrhosis or other liver diseases, stroke, breast cancer, andrenal failure (7, 22, 23). S. agalactiae colonizes the lower gas-trointestinal and genitourinary tracts of 30 to 50% of healthyadults (26), and an estimated 20 to 30% of all pregnant womenare carriers (25). S. agalactiae can be isolated from vaginal orrectal swabs, and prenatal screening for colonization of preg-nant women is recommended (27). The objective of this studywas to analyze the epidemiology of GBS in East African adultsby using multilocus sequence typing (MLST) (13) and capsularserotyping (10), with a focus on investigating possible correla-tions between clonal complexes of GBS in invasive or coloniz-ing isolates.The Aga Khan University Hospital in Nairobi (AKUH,N) isa tertiary care, referral university hospital in Nairobi, Kenya.GBS were cultivated at the Division of Microbiology ofAKUH,N from specimens derived from both inpatients andoutpatients between January 2007 and June 2010. Swabs fromoutreach collection points were transported in Stuart’s trans-port medium to the hospital. Blood cultures were routinelyperformed using Plus-aerobic/F, Plus-anaerobic/F, and PedsPlus Bactec 9120 media (Becton Dickinson) in combinationwith commercially available biphasic culture medium bottles(Roche Diagnostics, France). Single GBS colonies were pickedfrom 5% sheep blood agar plates (Oxoid, United Kingdom),and identification of GBS was performed using colony mor-phology, Gram staining, CAMP test, the API Streptococcusidentification kit (Bio-Merieux, France), and the latex aggluti-nation test using a bacterial meningitis kit with specific groupB latex (Wellcogen kit from Remel Europe Ltd.). Antimicro-bial susceptibility testing was performed, and results were in-terpreted according to the guidelines of the Clinical andLaboratory Standards Institute (CLSI) (5). Streptococcus pneu-moniae ATCC 49619 was used for quality control. All GBSisolates were found to be susceptible to penicillin and cepha-losporin. GBS isolates analyzed here were selected accordingto availability of stocked isolates and accessibility of patientfiles. One isolate per patient was included in this study. Clinicalinformation was obtained by review of patient files. Ethicalclearance for this study was obtained from the Ethikkommis-sion beider Basel (EKBB) and the AKUH scientific and ethicalcommittee.For MLST, GBS isolates were grown overnight at 37°C onplates with Columbia agar enriched with 5% sheep blood(Oxoid). Genomic DNA extraction, MLST PCR amplification,and sequencing of seven housekeeping genes were performedas described previously (13). Novel alleles of the housekeepinggenes were amplified and sequenced twice and submitted toNicola Jones, Oxford, United Kingdom, for allele assignment.The sequence types (STs) were determined by the S. agalactiaeMLST website (http://pubmlst.org/sagalactiae/) (12). The neigh-bor-joining tree was generated from the allelic profiles ob-tained using Start2 (11). The multiplex PCR assay for identi-fication of capsular serotypes was performed as describedpreviously (10). To assess if any clonal complex is associatedwith colonizing or invasive isolates, the distribution of eachclonal complex was examined against random distribution byusing the Pearson chi-square test. Association betweenclonal complex and capsular serotype distributions was as-sessed by using Fisher’s exact test. Statistical analyses wereperformed using STATA version 10 (Stata Corporation,College Station, TX).Ninety-five colonizing GBS isolates were derived from vag-inal swabs collected during routine prenatal screening of preg-nant women. Out of the 74 invasive isolates, 40 (54%) werederived from urine, 10 (13.5%) from blood, 8 (10.8%) frompus, and 16 (21.6%) from other body locations. Thirty-seven(50%) of the GBS from urine samples originated from women.* Corresponding author. Mailing address: Aga Khan UniversityHospital, Nairobi, 3rd Parklands Avenue, Nairobi, Kenya. Phone: 254-20-3662226. Fax: 254-20-3741749. E-mail: gunturu.revathi@aku.edu.‡ These authors contributed equally. Published ahead of print on 24 August 2011.3652 on November 9, 2012 by guesthttp://jcm.asm.org/Downloaded from Twenty-one of these nonpregnant female patients had re-peated attacks of symptomatic urinary tract infection with sig-nificant pyuria and heavy vaginal colonization. Nine patientswere antenatal symptomatic women with significant pyuria andbacteriuria. Two male patients with chronic GBS urinary tractinfection suffered from prostatic hypertrophy. One elderlymale patient suffered from catheter-associated GBS urinarytract infection due to neurogenic bladder. This patient devel-oped Pseudomonas aeruginosa bacteriuria after GBS subsidedunder antibiotic treatment (8, 21). Five of the patients withpositive blood cultures were elderly adults with comorbidities,including diabetes, renal failure, cardiac asthma, and malig-nancies with primary GBS bacteremia. One adult male devel-oped GBS bacteremia secondary to peritonitis and later pro-gressed to renal failure subsequent to toxic shock. Out of the 8pus isolates, 4 came from cases of osteomyelitis and one eachcame from a diabetic foot, liver abscess, pelvic abscess aspirate,and joint fluid. A fat tissue isolate was cultured from postmor-tem samples from an elderly woman. Five isolates from ure-thral swabs came from patients with nongonococcal urethritis(16).In total, 22 STs could be distinguished in this collection, withthe colonizing and invasive GBS isolates grouped into 16 STsand 19 STs, respectively (Table 1). Isolates were grouped intoclonal complexes (CCs) by eBURST, relaxing the group defi-nition to six out of seven shared alleles. Thirteen STs wereclustered in five CCs, namely, CC23, CC19, CC17, CC10, andCC1 (Fig. 1). The most prevalent CC was CC23 (27.2%),followed by CC17 (21.3%), CC10 (16.6%), CC19 (13.6%), andCC1 (12.4%). Eighty-eight percent of the isolates could begrouped into one of these five CCs. These five CCs are alsohighly prevalent in other GBS collections representing eitherworldwide collections (24) or investigations performed in Swe-den (17), Italy (9), England (14), the United States (3), theCentral African Republic, and Senegal (4) as well as Israel (1).ST-26, which represented 15% of all identified GBS isolates inthe Central African Republic and Senegal (4), was found threetimes in our collection only. Singletons identified were ST-3,ST-4, ST-24, ST-26 ST-103, ST-327, ST-328, ST-485, and ST-486 (Table 1). Five novel STs named ST-484, ST-485, ST-486,ST-492, and ST-501 were detected. ST-485 is a double-locusvariant and ST-486 is a single-locus variant of ST-103. ST-484is a single-locus variant of ST-17, while ST-492 and ST-501 aresingle-locus variants of ST-23. All novel STs were found atleast once in clinical isolates, and ST-484 and ST-486 wereadditionally found in colonizing isolates. The most prevalentnovel ST was ST-484, found in 6 case isolates and 2 colonizingisolates. ST-4, ST-103, ST-327, ST-485, ST-492, and ST-501were found exclusively in invasive GBS isolates. ST-3, ST-24,and ST-8 could be detected only among the colonizing GBSTABLE 1. Overview of the GBS sequence types and capsular serotypes obtained in the current studyCC and/or STNo. of isolates Capsular serotype(s) (no. of isolates)Total Colonizing Invasive Total Colonizing InvasiveCC1aST-1 16 9 7 V (15), Ia (1) V (9) V (6), Ia (1)ST-2 5 4 1 II (4), V (1) II (4) V (1)CC10aST-8 5 5 0 Ib (5) Ib (5)ST-10 20 10 10 Ib (18), II (2) Ib (9), II (1) Ib (9), II (1)ST-12 3 1 2 V (2), Ib (1) V (1) V (1), Ib (1)CC17aST-17 28 20 8 III (28) III (20) III (8)ST-484 8 2 6 III (8) III (2) III (6)CC19aST-19 8 6 2 III (5), V (3) III (5), V (1) V (2)ST-28 6 3 3 II (6) II (3) II (3)ST-182 9 4 5 III (9) III (4) III (5)CC23aST-23 44 24 20 Ia (42), III (2) Ia (24) Ia (18), III (2)ST-492 1 0 1 Ia (1) Ia (1)ST-501 1 0 1 Ia (1) Ia (1)SingletonsST-3 1 1 0 IV (1) IV (1)ST-4 1 0 1 Ia (1) Ia (1)ST-24 1 1 0 Ia (1) Ia (1)ST-26 3 2 1 V (3) V (2) V (1)ST-103 1 0 1 Ia (1) Ia (1)ST-327 1 0 1 V (1) V (1)ST-328 4 2 2 V (4) V (2) V (2)ST-485 1 0 1 Ia (1) Ia (1)ST-486 2 1 1 Ia (2) Ia (1) Ia (1)a Fisher’s exact test P value, 0.000 (comparison among CCs).VOL. 49, 2011 NOTES 3653 on November 9, 2012 by guesthttp://jcm.asm.org/Downloaded from bacteria (Table 1). Interestingly, out of the 10 isolates obtainedfrom blood, the novel ST-484, ST-485, ST-486, and ST-492represented 30% (Table 2), supporting their high invasive po-tential. The possible relationship between invasive or coloniz-ing GBS and the five CCs was analyzed (Table 2). No statisti-cally significant correlation between any of the five CCs andinvasive or colonizing GBS isolates could be described by usingPearson’s chi-square test (P  0.926). However, here we com-pare colonizing isolates from pregnant woman with invasiveisolates from nonpregnant adults.The capsular serotypes detected here included Ia, Ib, II, III,IV, and V. The most common serotype was serotype III(30.8%), followed by serotype Ia (30.2%), serotype V (17.2%),serotype Ib (14.2%), and serotype II (7.1%); one isolate was ofserotype IV (0.6%). Out of the 44 isolates identified as ST-23,42 were serotype Ia. All ST-17 isolates carried serotype III, 18FIG. 1. Neighbor-joining tree demonstrating the relationship between the 22 STs of the GBS identified in the current study. Thephylogenetic tree is based on the alleles of the seven housekeeping alleles sequenced and was constructed using the Start2 program(http://pubmlst.org/software/analysis/start2/). The scale bar indicates distance as calculated by the number of mismatched loci.TABLE 2. Sequence types and capsular serotypes of the GBS clinical isolates according to location of originCC and/or STNo. of isolates Capsular serotype(s) (no. of isolates)Total Urine Blood Pus Other Total Urine Blood Pus OtherCC1aST-1 7 4 1 2 V (6), Ia (1) V (4) Ia (1) V (2)ST-2 1 1 V (1) V (1)CC10aST-10 10 5 1 4 Ib (9), II (1) Ib (5) II (1) Ib (4)ST-12 2 1 1 V (1), Ib (1) V (1) Ib (1)CC17aST-17 8 7 1 III (8) III (7) III (1)ST-484 6 3 2 1 III (6) III (3) III (2) III (1)CC19aST-19 2 2 V (2) V (2)ST-28 3 3 II (3) II (3)ST-182 5 2 2 1 III (5) III (2) III (2) III (1)CC23aST-23 20 10 2 3 5 Ia (18), III (2) Ia (9), III (1) Ia (2) Ia (2), III (1) Ia (5)ST-492 1 1 Ia (1) Ia (1)ST-501 1 1 Ia (1) Ia (1)SingletonsST-4 1 1 Ia (1) Ia (1)ST-26 1 1 V (1) V (1)ST-103 1 1 Ia (1) Ia (1)ST-327 1 1 V (1) V (1)ST-328 2 2 V (2) V (2)ST-485 1 1 Ia (1) Ia (1)ST-486 1 1 Ia (1) Ia (1)a Pearson’s chi-square test P value, 0.926 (comparison among CCs).3654 NOTES J. CLIN. MICROBIOL. on November 9, 2012 by guesthttp://jcm.asm.org/Downloaded from out of 20 ST-10 isolates had serotype Ib, 15 out of 16 ST-1isolates expressed V, and all ST-484 and ST-182 isolates hadserotype III. Hence, a tight relationship between distinct cap-sular serotypes and the CC could be observed (Fisher’s exacttest P  0.000) (Table 1). Of note, invasive GBS typed asST-23, ST-12, and ST-1 showed a higher variability in theircapsular serotypes than colonizing isolates (Table 1). In sum-mary, our data show that the population structure of GBSisolates in East African adults is similar to the GBS populationstructure reported in industrialized countries. A number ofnovel STs that share alleles with housekeeping genes of wide-spread known STs were detected.We are thankful to the team in the Division of Microbiology, De-partment of Pathology, at the AKUH,N. The excellent assistance ren-dered by Patricia Shivachi with data management is gratefully ac-knowledged. 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Characterization of invasive and colonizing isolates of Streptococcus agalactiae in East African adults

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