Abstract
Group A streptococcal (GAS) pharyngeal colonization rates were determined among 1061 asymptomatic students in Hawaii and American Samoa where acute rheumatic fever rates are high. All GAS isolates were emm sequence typed. Although pharyngeal colonization rates were low in Hawaii (3.4%), Pacific Islander children had significantly higher colonization rates (5.7% versus 1.2% in other ethnic groups, p<0.05). The colonization rate was higher in American Samoa (13%). Few emm types that were infrequently observed in symptomatic infections in Hawaii were repeatedly identified in both sites. These emm types were previously described among asymptomatic children suggesting a type-specific association with pharyngeal colonization.
Keywords: Streptococcus pyogenes, epidemiology, asymptomatic colonization, emm typing, Pacific Islander
Group A streptococci (GAS) are responsible for a wide array of illnesses and sequelae such as acute rheumatic fever (ARF) [1]. The annual incidence rates of ARF have been endemically high in Hawaii, specifically among the Pacific Islander (PI) including Samoan children [2–4]. ARF is also reported to be common in American Samoa (AS) (The Central Intelligence Agency. The World Fact Book at https://www.cia.gov/library/publications/theworldfactbook/index.html) [5–7]. The reasons for high ARF rates among Samoans are not fully explained. Previously, we have shown infrequent continental US emm/M types among isolates identified from symptomatic patients in Hawaii (CDC Streptococcus pyogenes database at http://www.cdc.gov/ncidod/biotech/strep/strepindex.htm) [8–11]. Additionally, GAS isolates temporally associated with ARF in Hawaii belonged to types that have not been classically associated with ARF [3].
These results prompted us to determine the colonization rates in school children known to be the major reservoirs of GAS [12]. Although a wide range of colonization rates, from 2.2% to 29%, have been reported, emm typing and possible variations in colonization rates between different ethnic groups were inadequately addressed [13–18].
The Oahu, Hawaii study was conducted in April and May 2003. The Pago Pago, AS study was conducted in May 2006. Necessary approvals from the institutional review boards were obtained. Eleven Oahu schools, representing a cross section of ethnic and socioeconomic groups, and two Pago Pago, AS schools were selected. In Oahu, several ethnic groups including the Native Hawaiians and PI groups (Samoans, Tongans, Marshallese and Micronesians) live together.
Throat swabs were taken from all students, excluding those with symptomatic or clinical pharyngitis [19]. Isolates were identified as Streptococcus pyogenes by standard methodology and characterized by emm sequencing (http://www.cdc.gov/ncidod/biotech/strep/protocol_emm-type.htm). Emm types were then compared to a convenience sample of GAS isolates identified in Oahu [20]. Non-group A beta-hemolytic streptococci were identified using the API 20 Strep identification system (BioMerieux, St Louis, MO, USA).
SAS software and the variance formula in a clustered design was used to determine the colonization rates (α of 0.05 (two-sided) and a power of 80%). Multivariate analysis was used for prevalence data by school and ethnicity in Hawaii.
Throat cultures of 955 asymptomatic Oahu students (Grades K-12) were obtained. Of these, 488 (51.1%) were males. The mean age was 13.2 (SD 2.7) years. 457 (47.8%) students were of PI ethnicity. GAS was identified in 32 students (3.4%). The pharyngeal prevalence was 5.7% (26/457) in PI children vs. 1.2% (6/498) in other ethnic groups (p<0.05). Sixteen distinct emm types were found and six belonged to infrequent continental US types: emm 78, 95, 110, 116, st11014, stkNB4 (http://www.cdc.gov/ncidod/biotech/strep/strepindex.htm) [11]. Thirteen students had throat swabs repeated (minimum 18 month interval from the first culture). Two PI students had positive GAS swabs (Table 1).
Table 1.
Emm types in children from Oahu. The ethnicity that the student and family mostly identified with is marked as “*”
| School number-study number (32 students) | Ethnicity | Emm types | Follow-up visit (13 students) |
|---|---|---|---|
| 01-0014 | PI | 41 | Not done |
| 01-0015 | PI*, C, F, W | 78 | BHS, not A, B, C, G |
| 01-0020 | PI | 12 | Not done |
| 01-0032 | PI | 12 | Not done |
| 01-0034 | PI | ST11014 | NTF |
| 01-0048 | PI*, F | 11 | Not done |
| 01-0050 | PI*, W | 110 | BHS, not A, B, C, G |
| 01-0056 | PI*, C, W | 75 | BHS, not A, B, C, G |
| 01-0058 | PI*, F, W | 58 | Not done |
| 01-0090 | PI*, F, W | 78 | GAS, emm3 |
| 01-0100 | PI*, C, W | 77 | GAS, emm 77 |
| 01-0101 | PI*, C, F | 1 | Not done |
| 02-0048 | PI*, C | 12 | Not done |
| 02-0079 | PI | 95 | Not done |
| 03-0075 | PI*, F, W | 3 | Not done |
| 03-0094 | C | 58 | Not done |
| 03-0098 | PI*, C, F, W | 58 | Not done |
| 04-0027 | PI*, F | 3 | NTF |
| 05-0004 | PI*, C, F, W | 12 | Not done |
| 05-0037 | PI*, C, F | 11 | Not done |
| 05-0039 | W | 12 | NTF |
| 05-0054 | PI*, C | 116 | NTF |
| 05-0073 | PI*, W | 1 | Not done |
| 05-0085 | PI*, C, F | 92 | Not done |
| 06-0021 | A | 12 | Not done |
| 06-0062 | W | 28 | NTF |
| 07-0003 | W | 12 | GGS |
| 08-0013 | PI*, C, F | 3 | Not done |
| 09-0204 | W*, PI | 1 | NTF |
| 10-0034 | PI*, C, W | 77 | GGS |
| 11-0001 | F*, C | 1 | Not done |
BHS: Beta-hemolytic streptococci
C: Chinese
F: Filipino
GGS: Group G streptococcus
NTF: Normal throat flora
PI: Pacific Islander (including Samoans, Tongans, Micronesians and Marshallese)
W: White
Throat cultures were done in 106 Samoan students in AS, mean age 11.62 (SD:3.4). Fourteen (13%) were positive for GAS. Twelve emm types were found (Table 2). The colonization rate was higher in AS when compared with the Oahu study that included mixed ethnic groups (p-value: 0.0002). Eleven (10%) swabs were positive for group G streptococci. Thirteen students were retested and four (31%) had positive throat cultures. Three of these types were new and different from the first visit (emm types 22, 44, 74). Emm types 1, 92, 116 were identified in both Oahu and AS. The emm types identified in AS were identified in ~25% of Hawaiian isolates during the preceding six years [3, 20].
Table 2.
Emm types in children from Pago Pago.
| Student number (14 students) | First visit (May 2006, 14 students) | Second visit (September 2006) | Third visit (February 2007) |
|---|---|---|---|
| Emm types | Throat swab | Throat swab | |
| 1 | stNS 554 | GAS, emm stNS 554 | NTF |
| 2 | 92 | NTF | BHS, not A, B, C, G |
| 3 | 105 | GAS, emm 44 | Not done |
| 4 | 101 | GAS, emm 74 | BHS, not A, B, C, G |
| 5 | 56 | BHS, not A, B, C, G | Not done |
| 6 | 98 | GAS, emm 22 | BHS, not A, B, C, G |
| 7 | 25 | not isolated | GAS, emm 25 |
| 8 | 1 | GCS | Not done |
| 9 | 98 | NTF | Not done |
| 10 | 55 | BHS, not A, B, C, G | GAS, emm 89 |
| 11 | st 6030 | NTF | not isolated |
| 12 | 92 | GCS | not isolated |
| 13 | 108 | BHS, not A, B, C, G | Not done |
| 14 | 116 | Not done | Not done |
BHS: Beta-hemolytic streptococci
GCS: Group C streptococcus
GGS: Group G streptococcus
NTF: Normal throat flora
This is the first large scale study indicating ethnic differences for GAS pharyngeal colonization in Hawaii. It is unclear if high colonization rates would directly influence the transmission or sequelae rates. There may obviously be genetic bias toward increased carriage or ARF rates in these population groups that also have the highest ARF rates in Hawaii (9.5–12.4/100,000 population) [3, 21]. The AS survey indicated even higher rates of pharyngeal colonization. No similar association of high colonization rates among regions disproportionately affected with ARF has been reported in multiethnic populations [14, 18, 22]. GAS colonization is dynamic and the presence of certain emm types in children could presumably affect the rest of the population and can spread in and out of specific populations [23]. Similarly, acquisition and spread of GAS in Samoans who live in close-knit populations in Hawaii and frequently travel back and forth from Samoa are possible in the isolated island settings of this study. The different study periods limited a direct comparison between the two sites, however, a high number of emm types were identified in both Hawaii and AS and few of these types were identified in both studies. Emm types 1, 3, 12, 28, 75 and 77 were similarly identified in recent studies [14, 22]. This type correlation among asymptomatic children to other geographic areas is significant. A similar correlation was not observed among isolates identified from patients with clinical illness in Hawaii. On the contrary, we had identified a higher number of infrequent continental U.S. emm types in patients with clinical disease (http://www.cdc.gov/ncidod/biotech/strep/strepindex.htm) [3, 8, 10, 11, 20].
Unlike ARF and invasive GAS disease, emm types associated with asymptomatic colonization or carrier state are not well-studied. Identification of similar emm types from different geographic regions with different GAS disease epidemiology may be explained by the biologic affinity of few, unique emm types to simply cause asymptomatic colonization [14, 22, 24]. Interestingly, some of the emm types, such as emm 44/61, 92 and 98, identified among asymptomatic students were also identified from nine ARF patients in Hawaii (3 and unpublished data). The higher colonization rates and emm type similarities would not completely explain the disproportionate disease burden in Samoans but rather point to additional bacteriologic factors.
Overall GAS colonization rates and GGS colonization of asymptomatic and ARF-affected Samoan children are worth mentioning. In some previous studies, colonization rates were reported to be high (up to 25%) [13, 14]. GAS colonization rates in Oahu were similar to data reported from clinical practices and therapeutic trials [15, 17, 23]. Higher rates of colonization were seen only in the population groups affected with ARF. These children also showed higher colonization rates with GGS and other beta hemolytic non-group A streptococci. The 10% GGS colonization in AS was much higher than some of the previously reported colonization rates but was similar to the rates reported in Australian Aborigines who are also disproportionately affected by ARF [15, 16, 25, 26].
The colonizing emm types in a community with high rates of ARF and other streptococcal infections need to be considered in designing candidate vaccines made to utilize type-specific immunity. Based on our sample, such a candidate vaccine would provide coverage for only 57% of isolates (27), which is higher than we have observed in clinical infections but still probably not optimal.
In summary, we observed higher GAS colonization rates among Pacific Islander children in Hawaii and American Samoa. We have also identified emm types that are uncommon in Hawaii but that were previously described among asymptomatic streptococcal carriers suggesting a type-specific association with pharyngeal colonization.
Acknowledgments
Transparency declaration
This publication was made possible by Grant Number 5 P20R018727 from the National Center for Research Resources (NCRR), grant number P20RR11091 from Research Centers in Minority Institutions, both components of the National Institutes of Health (NIH), and grant number 0265237Z from American Heart Association. Preliminary study findings were presented at the 43rd Annual Meeting of the Infectious Diseases Society of America, San Francisco, CA, October 2005 (abstract 1085) and at the Annual Pediatric Academic Societies Meeting in Toronto, Canada, May 2007 (abstract 752221).
Footnotes
Transparency Declaration
All authors declare no conflicts of interest.
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