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treptococcal Upper Respiratory Tract Infectionsnd Psychosocial Stress Predict Future Tic andbsessive-Compulsive Symptom Severity inhildren and Adolescents with Tourette Syndromend Obsessive-Compulsive Disorder

aiqun Lin, Kyle A. Williams, Liliya Katsovich, Diane B. Findley, Heidi Grantz, Paul J. Lombroso,obert A. King, Debra E. Bessen, Dwight Johnson, Edward L. Kaplan, Angeli Landeros-Weisenberger,eping Zhang, and James F. Leckman

ackground: One goal of this prospective longitudinal study was to identify new group A beta-hemolytic streptococcal infections (GABHS)n children and adolescents with Tourette syndrome (TS) and/or obsessive-compulsive disorder (OCD) compared with healthy controlubjects. We then examined the power of GABHS infections and measures of psychosocial stress to predict future tic, obsessive-compulsiveOC), and depressive symptom severity.

ethods: Consecutive ratings of tic, OC, and depressive symptom severity were obtained for 45 cases and 41 matched control subjectsver a 2-year period. Clinical raters were blinded to the results of laboratory tests. Laboratory personnel were blinded to case or control statusnd clinical ratings. Structural equation modeling for unbalanced repeated measures was used to assess the sequence of new GABHS

nfections and psychosocial stress and their impact on future symptom severity.

esults: Increases in tic and OC symptom severity did not occur after every new GABHS infection. However, the structural equation modelound that these newly diagnosed infections were predictive of modest increases in future tic and OC symptom severity but did not predictuture depressive symptom severity. In addition, the inclusion of new infections in the model greatly enhanced, by a factor of three, theower of psychosocial stress in predicting future tic and OC symptom severity.

onclusions: Our data suggest that a minority of children with TS and early-onset OCD were sensitive to antecedent GABHS infections.

hese infections also enhanced the predictive power of current psychosocial stress on future tic and OC symptom severity.

ey Words: Attention-deficit/hyperactivity disorder, obsessive-ompulsive disorder � Tourette syndrome � tic � group A betaemolytic streptococcal upper respiratory tract infections, majorepressive disorder (MDD), PANDAS, structural equation modeling

ourette syndrome (TS) and pediatric onset obsessive-compulsive disorder (OCD) are etiologically related, chronic,familial neuropsychiatric disorders that affect as many as .3%

o 3% of the pediatric population (1,2). They are chronic disordershat can be associated with impairment and disability.

The etiologies of these disorders are unknown. It has beenypothesized that some susceptible individuals develop theseymptoms because of postinfectious autoimmune processes. Infec-ions with group A beta-hemolytic streptococci (GABHS) have beenypothesized to be responsible. Swedo et al. (3) identified thisubgroup with the acronym PANDAS (pediatric autoimmune neu-

rom the Department of Epidemiology and Public Health (HL, HZ) and ChildStudy Center (KAW, LK, DBF, HG, PJL, RAK, AL-W, HZ, JFL), Yale Center forClinical Investigation, Yale University School of Medicine, New Haven,Connecticut; Department of Microbiology and Immunology (DEB), NewYork Medical College, Valhalla, New York; World Health Organization (DJ,ELK), Streptococcal Reference Laboratory, Department of Pediatrics,University of Minnesota School of Medicine, Minneapolis, Minnesota.

ddress correspondence to James Leckman, M.D., Child Study Center, YaleUniversity School of Medicine, New Haven CT 06520-7900; E-mail:james.leckman@yale.edu.

eceived Aug 15, 2008; revised Jun 30, 2009; accepted Aug 8, 2009.

006-3223/10/$36.00oi:10.1016/j.biopsych.2009.08.020

ropsychiatric disorders associated with streptococcal infections). Inaddition to OCD, TS, and tic disorders (3–5), a specific link betweenattention-deficit/hyperactivity disorder (ADHD) and GABHS hasbeen hypothesized but not proven (3,5–7).

Many studies have emphasized the importance of psychoso-cial stress in mediating the progression of autoimmune diseases(8–11). Earlier cross-sectional and longitudinal studies of TS andearly-onset OCD have consistently suggested that these disordersare sensitive to stress and show a heightened stress response(12–14). Psychosocial stress has also been shown to be animportant factor in the onset and course of ADHD and majordepressive disorder (MDD) (15–17).

The goal of this blinded prospective longitudinal study was toexamine simultaneously the impact of both newly diagnosedGABHS infections and psychosocial stress in children and ado-lescents with TS and/or OCD. We had previously shown in thissame cohort the impact of psychosocial stress on future tic,obsessive-compulsive (OC), and depressive symptom severity(12). Our a priori hypothesis was that both new GABHS infec-tions and psychosocial stress would influence future fluctuationsin tic, OC, and depressive symptom severity. We applied astructural equation modeling framework with latent-time varyingconstructs to maximize our ability to examine this hypothesis.

Methods and Materials

SubjectsThe 86 subjects in this study were aged 7 to 17 years. Forty-five

case and 41 healthy control subjects were recruited. During a 2-year

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tudy period, participants were monitored, on average, monthlyy telephone and thrice-yearly with in-person visits. All patientsere followed at the Yale Child Study Center (April 1999–eptember 2004) (18–20). Community control subjects wereecruited from a list of 10,000 names purchased from a telemar-eting company. Control subjects were group-matched for age,ex, socioeconomic status, and season of the year at entry intohe study. Control subjects were selected only if there was noifetime personal history, for the patient or any first-degreeelative of a DSM-IV diagnosis of a chronic tic disorder, TS, OCD,r ADHD. No ethnic or racial groups were excluded. Informationn past GABHS infections was not used in making decisionsegarding participation.

Exclusion criteria for both groups included an IQ �75; aerious medical or neurological illness; major sensory handicapsblindness, deafness); head trauma resulting in loss of conscious-ess; current (past 6 months) psychiatric disorder that couldnterfere with participation, such as MDD; psychosis; or aervasive developmental disorder.

The protocol was approved by the internal review boards athe Yale University and the central streptococcal laboratory at theniversity of Minnesota. All parents provided written informedonsent after the study was described to them in detail. Aeparate assent form was used to ensure the informed participa-ion of the child and adolescent subjects.

linical AssessmentsWhen a family entered the study, information concerning the

atient was collected in a two-stage process (14,18–21). Initiallyhe families, in conjunction with experienced clinicians, com-leted the Yale Global Tic Severity Scale (YGTSS) (22) and thehildren’s Yale-Brown Obsessive Compulsive Scale (CY-BOCS)23) using both a self- and family-reports. Depression wasssessed with the children’s Depression Rating Scale-RevisedCDRS-R) (24). Anxiety was measured with the Revised Chil-ren’s Manifest Anxiety Scale (RCMAS) (25).

Comorbid psychiatric diagnoses, including ADHD, wereade by three expert clinicians (J.F.L., R.A.K., and P.J.L.) after

eviewing all available information including data from thechedule for Affective Disorders and Schizophrenia for School-ge Children (26). The same diagnostic panel used the PANDASriteria (3) to classify patients as possible, probable, definiteANDAS, or non-PANDAS cases. These criteria require theresence of OCD and/or a chronic tic disorder, an age at onsetetween 3 years and the beginning of puberty, a clinical courseharacterized by the abrupt onset of symptoms or by a pattern oframatic recurrent symptom exacerbations and remissions, aemporal relationship between GABHS infections and the clinicalourse of illness, and neurologic abnormalities. Patients withANDAS diagnoses of “probable” or “definite” were classifiednto the PANDAS group.

In addition, measurements of psychosocial stress were madesing parent report (Parent Perceived Stress Scale [PSS-P]) (27),outh self-report (Daily Life Stressors Scale [DLSS]) (28), and alinician-rated measure of long-term contextual threat (Yale chil-ren’s Global Stress Index [YCGSI]) (19). Normal control subjectsere also assessed for their stress level for comparison with the

ubjects who had been diagnosed with TS and/or OCD (12).

ongitudinal EvaluationsTic and OC symptom severities were rated by an expert rater

t the approximately monthly telephone contacts using the tic

ortion of the YGTSS and the CY-BOCS. Clinical raters were

blinded to the results of laboratory tests. In-person visits oc-curred approximately every 4 months for additional monitoringof clinical severity and collection of biological specimens. ThePSS-P ratings were made at each telephone and each in-personencounter, and the DLSS and the clinician-rated YCGSI wereobtained every 4 months. Data collected from this cohort of casesand control subjects concerning the effects of longitudinal psy-chosocial stress on future ratings of tic, OC, and depressivesymptom severity using an identical analytic strategy has beenpreviously reported (12).

As previously described in detail, tic and/or OC symptomexacerbations were identified using a two-stage algorithm (18). Ifan exacerbation occurred, the patient was immediately reevalu-ated and scheduled for two in-person visits when additionalbiological specimens were collected (an “exacerbation” visit wasscheduled as soon as possible, and a “follow-up” visit wasscheduled 2 months after the exacerbation visit). Most familiescompleted their regularly scheduled encounters. However, thenumber of encounters varied among the subjects from 4 to 26(median � 24). However, some of the intervals were greater than1 month. On eight occasions, the interval was greater than 100days. The mean time between consecutive staff–family encoun-ters, across all patients, was 32.3 days (SD � 11.9).

Biological SpecimensBlood samples and throat cultures were obtained approxi-

mately every 4 months. As described (20), following centrifuga-tion serum was stored in aliquots at �80°C. A total of 309 (cases)and 253 (control subjects) blood specimens were collected.Pharyngeal swabs were cultured on standard blood agar plates(20). Grouping of the streptococci was performed using a latexbead agglutination test (29). Throat cultures were obtained at 307patient and 250 control subject visits.

Laboratory personnel were blinded to case or control statusand clinical status. In addition, the results of the throat cultureswere not shared with the participants’ primary care clinicians. Weinformed the families before they enrolled and signed informedconsents and assents that we were not in a position to share theculture results with their primary care clinicians because thelaboratory performing the cultures (D.E.B.) was not certified toreport such results. If the family agreed to participate, we thentold them directly that if they had any concern about a possiblepharyngeal infection that they should contact their pediatricianimmediately and follow their treatment recommendations. Wealso sent letters to each of the clinicians and had a pediatric nursepractitioner visit each of the practices. They were encouraged tofollow whatever their standard procedures were and to treat thechild with antibiotics, if indicated. We also shared with them thestandard practice guidelines that were in place at that time (30).

GABHS InfectionsSerum obtained from each blood sample was assessed for

antistreptolysin O (ASO) and antideoxyribonuclease B (ADB)titers with standard methods (29,31,32). Table 1 presents theDefinite and Possible criteria used to diagnose new GABHSinfections on the basis of titer changes and the culture results(33). Culture data from the enrollment visits that lacked theinformation required to determine whether new infection hadoccurred were not used in the analyses. Similarly, data from visitsthat occurred more than 100 days after a culture visit were notused in the analyses. As a result, in the patient group, 225 visits

were available for evaluation. The mean time interval from a

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ulture visit to the following rating encounter was 33.4 daysSD � 13.8; range � 4–99 days).

edication StatusEach subject’s medication history was monitored at each

ncounter. Information was recorded according to the use ofntibiotics as well as the class of psychotropic agents (adrenergicgonists, neuroleptics, serotonin reuptake inhibitors and otherntidepressants, mood stabilizers, psychostimulants, benzodiaz-pines, and other agents). Baseline psychotropic medicationtatus and changes in medication status, including the use ofntibiotics (in separate models) were included in the structuralquation modeling.

tatistical AnalysisThe baseline demographic and clinical data for cases and

ontrol subjects as well as PANDAS and non-PANDAS subjectsere compared using either t test or Fisher Exact Test. A Poisson

egression model was used to compare rates of infections in theffected subjects group, normal control subjects, and PANDASnd non-PANDAS subgroups. A similar model was used toxamine the rate of exacerbations across all cases and separatelyor the PANDAS and non-PANDAS cases. To assess whetherxacerbations were temporally associated with an antecedentABHS infection, we examined the observed number of “hits” inach group along with a 95% confidence interval for the trueean number of hits (given the fixed number of person-years of

ollow-up in this study) based on the assumption that the numberf hits follows a Poisson distribution (33). Next, we used aepeated-measures analysis of variance to determine whetherhere was any change in symptom ratings following a newlyiagnosed GABHS infection.

Finally, a structural equation modeling framework was em-loyed to explore the relationships between newly diagnosedABHS infections, stress, severity of tic and OC symptoms, andepression. All the coefficients in a structural equation modelere simultaneously estimated with the maximum likelihoodethod using the Stata add-on module GLLAMM (34,35) because

t accommodates different measurement schedules among mul-iple longitudinal measures. First, we incorporated newly diag-osed GABHS infections as a binary variable into the longitudinalath models. The timing of the newly diagnosed GABHS infec-ions (Definite or Possible) was based on the date of the second

able 1. Criteria to Determine Possible and Definite Streptococcalnfectionsa

ossible GABHS Infections Definite GABHS Infections

ither one of the following conditions(but not both) have to be met:

. A log10 SO or ADB titer increase of .2R. A new isolation of GABHS without a

corresponding rise in antibody titer.

Any of the following conditionshave to be met:

1. A rise in titer of .2 log in bothASO and ADB antibodies

OR2. A rise in titer of greater than .2

log in one or both antibodiesOR3. isolation of GABHS with a

corresponding rise in anantibody titer of �.2 log

ADB, antideoxyribonuclease B; ASO, antistreptolysin O; GABHS, group Aeta-hemolytic streptococci.

a The timing of the new diagnosed infections was set at the point whenhe titer change was detected.

f two consecutive blood samples that met the criteria presented

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in Table 1. All baseline data pertaining to GABHS infections wereexcluded. Separate runs were also performed comparing newlydiagnosed GABHS infections defined by Definite criteria versusDefinite or Possible criteria.

As in our earlier study (12), a latent time-varying stressconstruct was modeled to represent the underlying true stresslevel manifested by the three available longitudinal stress mea-sures: the YCGSI, the PSS-P, and the DLSS. Time lag effect wasstudied with mixed-effect submodels in a structural equationframework by using the nearest recorded past reading of alongitudinal measure as a covariate for a current reading. Theeffects of antecedent newly diagnosed GABHS infections andprior stress level on current tic, OC, and depressive symptomseverity were all explored. We also excluded all data points forwhich the interval was greater than 100 days (n � 8 events). Therelationships between newly diagnosed GABHS infections, psy-chosocial stress and tic, and OC and depressive symptomseverity were studied via the submodels within the structuralequation modeling framework. See Lin et al. (12) for additionaldetails. The correlations among various stress measures andtime points of a same measure in a same subject wereaccommodated by the stress construct that could be affectedby other covariates, including newly diagnosed GABHS infec-tions, in the same model (34,35).

Results

Description of Study CohortApart from sex, the cases and control subjects had similar

demographic characteristics (Table S1 in Supplement 1). Elevencases were given a presumptive diagnosis of PANDAS (probableor definite) based on their history (3). With the exception of sex,the PANDAS cases were similar to non-PANDAS cases andunaffected control subjects with respect to mean age, ethnicity,parent education, and age of symptom onset (Table S2 inSupplement 1).

At baseline, cases had higher ratings of anxiety and depres-sion as well as higher levels of psychosocial stress comparedwith the normal control subjects. PANDAS cases also had signif-icantly higher baseline levels of anxiety, depression, and psycho-social stress compared with the healthy control subjects group.No differences were noted comparing the PANDAS cases andnon-PANDAS cases on any of these variables (Table S2 inSupplement 1).

GABHS InfectionsThe PANDAS cases had the highest rate of Definite or Possible

GABHS infections (.40 per person per year) compared with thenon-PANDAS cases (.34 per person/year) or the unaffectedcontrol subjects (.35 per person/year; Table 2 and Table S3 inSupplement 1). The PANDAS group was also at an increased riskfor Definite GABHS infections when compared with the normalcontrol subjects or non-PANDAS patients. However, none ofthese differences were statistically significant (Table 3).

Antibiotic TreatmentOf the 35 Definite and 21 Possible new GABHS infections,

only five were treated with antibiotics. In the patient group, therewere three treated infections of 20 Definite and 10 Possibleinfections. In the control group, there were only two treatedinfections of 15 Definite and 11 Possible newly diagnosed

GABHS infections.

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elationship Between Newly Diagnosed GABHS Infectionsnd Tic and OC Symptom Exacerbations

First, we examined the rate of exacerbations (as a binaryes/no) across all cases and separately for the PANDAS andon-PANDAS cases. The number of exacerbations was highermong the PANDAS cases (.64 per person per year) than amonghe non-PANDAS cases (.40 per person per year), but thisifference was not significant. Next, we examined the number ofhits” (exacerbations that are temporally associated with eitherefinite or Definite plus Possible newly acquired GABHS infec-

ions) for all cases as well as for the PANDAS and Non-PANDASroups separately. These results were not significant (Tables4–S6 in Supplement 1).

Next, we examined whether a Definite or Possible newABHS infection was associated with either no change or an

mprovement in YGTSS and/or CY–BOCS scores at subjects’ nextncounter. There were 25 instances of a Definite or Possible newABHS infection. Seventeen (68%) of these encounters weressociated with either no change or an improvement in theirGTSS scores at their next encounter. A similar response wasbserved following the 200 visits identified as not having aossible or Definite new infection 139/200 (70%). Although theercentage of culture visits associated with a “worsening” of ticymptoms was similar following new infection (32%) and no newnfection (31%), the magnitude of the increase in YGTSS scoresas significantly greater in the infection group [mean increase of.1 � 6.0 vs. 4.6 � 3.3; F (1,17) � 3.28, p � .002]. A similar patternas seen with regard to changes in OC symptom severity such

hat 17 of the 25 Possible and Definite new infections (68%) and67 of 200 (84%) culture visits not representing new infections

able 2. GABHS Infections

roup

Period ofObservation

(months)

Definite GABHSInfections

(Infections perPerson per Year)

Definite � PossibleGABHS Infections

(Infections perPerson per Year)

ontrol Subjects 895 15 (.20) 26 (.35)ll Cases 1,020.5 20 (.24)a 30 (.35)b

ANDAS Cases 242 6 (.30) 8 (.40)on-PANDAS Cases 778.5 14 (.22) 22 (.34)

No significant differences were observed comparing the rates of infec-ions.

GABHS, group A beta-hemolytic streptococci; PANDAS, pediatric auto-mmune disorders associated with streptococcal infections.

aIn the longitudinal modeling, only 19 instances could be used becausen one instance the time interval between the visit at which a definitenfection was detected and the following visit was greater than 100 days.

bIn the longitudinal modeling only 25 instances could be used as fourossible infections were detected at the final visit of the study and for oneefinite infection the time interval between the visit at which the infectionas detected and the following visit was greater than 100 days.

Table 3. Relative Risks for GABHS Infections

Relative Risk for DefiniGABHS Infections

All Cases/Control Subjects 1.17PANDAS/Control Subjects 1.47PANDAS/Non-PANDAS 1.38Non-PANDAS/Control Subjects 1.07

None of the relative risks above were statistically sigCI, confidence interval; GABHS, group A beta-hemo

associated with streptococcal infections. No significant differe

were followed by either no change or by improvements in OCsymptoms. However, only the Definite plus Possible infectionswere modestly associated with an increase in OC symptomseverity [F (1,21) � 6.28, p � .02].

Pathway Analyses of the Relationship Among NewlyDiagnosed GABHS Infections, Measures of PsychosocialStress, and Future Ratings of Symptom Severity

Pathway analyses incorporating simultaneous modeling of allthe variables revealed that within the patient group, prioroccurrence of a Definite or Possible newly diagnosed GABHSinfection (p � .001) and past levels of psychosocial stress(controlling for the effects of antecedent depression) weresignificantly (p � .001) associated with a future worsening of ticseverity, as measured approximately 1 month later (Figure 1A).In the same model, increasing chronologic age (p � .001) wasassociated with tic improvement. The inclusion of subjects’ sex inthe model was not associated with changes in future tic severity.Although the predictive power of Definite newly diagnosedGABHS infections was modest, the inclusion of this variable inthe model increased the predictive power of the stress constructby a factor of almost three (from .13 to .37) (12). When that samemodel was run using both Definite and Possible newly diag-nosed GABHS infections, the effect of stress and GABHS infec-tions remained significant.

Pathway analyses also revealed that psychosocial stress andDefinite plus Possible (but not Definite alone) newly diagnosedGABHS infections were significant predictors of future OCsymptom severity (Figure 1B). Similar to the tic severity findings,the inclusion of Definite and Possible newly diagnosed GABHSinfections greatly increased, by a factor of more than three, thepower of psychosocial stress to predict future OC symptomseverity (12). Current OCD severity was also a statisticallysignificant predictor of future depressive symptoms.

Finally, together with depressive symptom severity, the Def-inite or Possible newly diagnosed GABHS infections greatlyincreased the current psychosocial stress level (Figure 1C).However, psychosocial stress, but not newly diagnosed GABHSinfections (Definite or Possible criteria), was associated with afuture worsening of depressive symptom severity.

Neither the severity of anxiety symptoms recorded over thecourse of the study nor a diagnosis of ADHD at entry wasstatistically related to the severity of tics or OC symptomsrecorded during the study period (data not shown). Likewise,when data concerning changes in a subject’s antibiotic or med-ication status were included as a predictor, there was no influ-ence on the relative significance of the directional findingsdepicted in Figure 1A–1C.

In an exploratory analysis, we also ran the model using justthe data from the 11 PANDAS cases. The effect of Definite orDefinite plus Possible newly diagnosed GABHS infections on

5% CIRelative Risk for Definite � Possible

GABHS Infections 95% CI

0, 2.28 1.01 .60, 1.717, 3.8 1.13 .51, 2.513, 3.58 1.17 .52, 2.622, 2.22 .97 .55, 1.71

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uture tic and OC symptom severity was even more significant inhe 11 PANDAS cases than in all the patients (data not shown).

iscussion

During this blinded, prospective longitudinal study, periodsf tic and OC symptom worsening were independently associ-ted with antecedent newly diagnosed GABHS infections as wells higher levels of antecedent psychosocial stress. Comparedith the earlier reports from the same sample (12), two novel

indings appear. First, the impact of psychosocial stress istronger when antecedent newly diagnosed GABHS infectionsre also considered in the model (Figure 1A and 1B). Second, inhis analysis using structural equation modeling, we found aignificant but modest effect of newly diagnosed GABHS infec-ions on future worsening of tic and OC symptom severity whenll the cases were evaluated. This was also the case when wexamined just the 11 PANDAS cases. However, the number ofutative PANDAS cases is too small to allow any firm conclu-ions.

A recently completed prospective longitudinal study thatncluded 40 matched pairs, PANDAS cases versus non-PANDASS/OCD cases, found that many of the tic and OC symptomxacerbations were not associated with newly diagnosed GABHS

Stress Construct(SCT)

PSS-P DLSSYCGSI

Time-lagged Depression (CDRS-RT-1)

Future Tic severity

(YGTSST+1)

Age-0.33* (<.001)

0.20* (.045)

0.37* (0.04)

1* (<.001) 1.29* (<.001)0.59* (<.001)

GABHS Infections

0.41* (.001)

A

Time–laggedStress Construct

(SCT-1)

PSS-P DLSSYCGSI

Future OC severity

(CY-BOCST)

Time-led Depression (CDRS-RT+1)

1* (<.001) 1.26* (<.001)0.64* (<.001)

.44* (.011).84*(<.001)

GABHS Infections

.22* (.043)

B

Stress Construct (SCT)

Time-led Depression (CDRS-RT+1)

Time-lagged Depression (CDRS-RT-1)

0.29* (<.001) 0.84* (<.001)

1* (<.001)0.89* (<.001)0.82* (0.009)

Time-lagged GABHS Infections

0.55* (<.001)

C

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infections and that most of the newly diagnosed GABHS infec-tions did not lead to an exacerbation (33). However, the potentialconfounding effects of antibiotics were not reported. Indeed, inthat study, of 25 Definite new infections, 11 (44%) were treatedand a similar rate was found for Possible new infections (31infections of which 13 (42%) were treated). These rates are 4times higher than in our study.

Although the actual rates of newly diagnosed GABHS infec-tions in our study were not significantly different across groups,

4™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™™Figure 1. Pathway analyses of the relationships between longitudinallycollected measures of newly diagnosed group A beta-hemolytic streptococ-cal (GABHS) infections, psychosocial stress and tic, obsessive-compulsive,and depressive symptom severity. Squares depict measured variables, andcircles represent latent variables. The standardized coefficients representthe size of the effects, and the p values in parentheses represent the signif-icance of these effects because of the Wald test. An effect is deemed to bestatistically significant if the p value for the Wald test for each of the �s, �s,and s is smaller than .05 (see text). The standardized coefficients are theproportion of 1.0 SD of the predicted variable that can be explained by 1.0SD change in the predicting variable. However, because GABHS infection isan event with a binary outcome for which analysis of SD is not appropriate,the GABHS coefficient was standardized using only the SD of the respectivesymptom severity measure. A solid line indicates a statistically significantpositive relationship; a dashed line indicates a significant inverse relation-ship. The thickness of the line reflects its relative significance. An arrowindicates a predictive relationship, and a line without an arrow head indi-cates the relative contributions to the latent variable of psychosocial stress.The stress construct (SC) is the latent construct derived from the Yale Chil-dren’s Global Stress Index (YCGSI), the Perceived Stress Scale—Parent (PSS-P),and the Daily Life Stress Scale (DLSS). The size of the coefficients associatedwith the SC indicate the relative contributions of the PSS-P, DLSS, and YCGSIto the latent stress construct. (A) Future tic severity. Future tic improvementwas associated with advancing chronological age. The standardized coeffi-cient of .33 means that following an increase of 2.7 years of age (1 SD of allmeasures of age in the patient population), we observed a reduction of 33%of 1.0 SD of all the YGTSS scores (.33 � 9.0 3.0). In addition, newlydiagnosed GABHS infections (defined by either the Definite or the Definiteplus Possible criteria) were associated with future worsening of tic severity.Because of the binary nature of the GABHS infection, the standardizedcoefficient of .41 (for Definite infections) means that the new infectionassociated with a 3.7-point increase of the YGTSS scores compared with theYGTSS score obtained approximately 1 month later (41% � 9.0 3.7). Inaddition, increases in past depressive symptoms (Children’s DepressionRating Scale-Revised, CDRS-RT�1) predict higher levels of current psychoso-cial stress, which in turn modestly, but significantly, predict increases infuture tic severity (Yale Global Tic Severity Scale, YGTSST�1). Increases incurrent depressive symptoms (Children’s Depression Rating Scale—Re-vised, CDRS-RT) are also modestly and independently predictive of increasesin future tic symptom severity (YGTSST�1). Prior tic symptom severity(YGTSST�1) does not affect current depressive symptom ratings, and pastYGTSS scores also do not significantly affect current stress levels (data notshown). In the case of future tic severity, the DLSS has the strongest predic-tive. The inclusion of newly diagnosed GABHS infections greatly increased(by a factor of almost three) the effect size of psychosocial stress to predictfuture tic symptom severity, with the standardized coefficient increasingfrom .13 to .37 (12). (B) Future obsessive-compulsive (OC) symptom severity.Future OC symptom worsening was associated with antecedent increases inSC and with Possible, but not Definite, newly acquired GABHS infections.Increases in Children’s Yale-Brown Obsessive Compulsive Scale (CY-BOCS)scores also predict future increases in depressive symptoms (CDRS-RT�1).The degree of increase in OC symptom severity following possible GABHSinfections was less than that seen in predicting future tic severity. However,the inclusion of newly diagnosed GABHS infections greatly increased by afactor of more than three the power of psychosocial stress to predict futureOC symptom severity as the standardized coefficient increased from .26 to.84 (12). (C) Future depressive symptom severity. Antecedent increases indepressive symptom severity with either Definite or Possible GABHS infec-tions predict higher levels of psychosocial stress. Current levels of psychos-

ocial stress, in turn, predict future depressive symptom severity.

PSS-P DLSSYCGSI

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H. Lin et al. BIOL PSYCHIATRY 2010;67:684–691 689

ur data add support to the suggestion that a subset of individualsith TS and/or OCD are at higher risk of developing GABHS

nfections relative to normal control subjects (4,5) or non-PANDASases (33). Two studies using large administrative databases haveoth reported that before initial tic disorder (4,5) or OCD (5)nset, a subset of patients was more likely to have a GABHSnfection compared with control subjects.

However, the design of our study—specifically, the timing ofulture collection for determining the presence of streptococcind sera for determining an immune response at the time ofymptom exacerbations—has the potential to bias our findingsn the direction of identifying a higher number of newly diag-osed GABHS infections in the patient group. This bias was notresent in a prospective longitudinal study by Kurlan et al. (33),ho also found that the PANDAS group had a higher rate ofefinite plus Probable GABHS infections than the non-pandasS/OCD cases, .43 versus .13 per person per year, respectively.he reasons underlying this increased risk require further study.hese cases may be at increased risk for GABHS infections, buthis increased risk may be unrelated to the etiology of theirondition.

Another important limitation of this study concerns the way inhich the timing of a newly diagnosed GABHS infection wasstimated. Because we relied on the timing of the routineollow-up visits when blood and culture specimens were ob-ained to diagnose new infections, the actual infection mightave occurred well before it was detected. More recent studies33) have involved more vigorous methods including routineisits every 3 months (rather than every 4 months), monthlyhroat cultures (rather than just collecting culture materials at theoutine and exacerbation visits), as well as collecting morelinical data to estimate more accurately the actual point whenhe infection occurred. Although the timing of the consecutivetaff–family encounters averaged 32 days, the interval on rareccasions was more than 100 days, and these data points werexcluded from the longitudinal modeling. In the final data set,4% of the staff–family encounters intervals were 45 days or less.

It has been postulated that GABHS infection must be thenitial autoimmune response-inciting event for PANDAS cases,ut subsequent symptom exacerbations may be triggered byrior infections with other infectious agents (5); a number ofther potential precipitating factors have been identified, includ-ng the common cold (36) and Mycoplasma pneumoniae (37–9). Future prospective longitudinal studies are needed to con-irm these findings and to clarify whether there is a commonnderlying immunologic mechanism at work.

The immunologic mechanisms that may underlie TS, OCD orANDAS, if any, remain in doubt. Increased activation of im-une responses has been suggested by changes in gene expres-

ion profiles of peripheral immune cells, relative frequency ofymphocyte subpopulations, and synthesis of immune effectorolecules. Increased activity of cell-mediated mechanisms is

uggested by the increased expression of genes controllingatural killer and cytotoxic T cells, increased plasma levels ofome proinflammatory cytokines that correlate with diseaseeverity, and increased synthesis of antineuronal antibodies20,40–56). However, this remains a controversial area of scienceecause several of these findings have not been independentlyeplicated (57–62). For example, a study using biological speci-ens from the earlier Kurlan et al. PANDAS study (33) failed to

eplicate earlier cytokine and antineuronal antibodies abnormal-ties in those patients in whom a newly diagnosed GABHS

nfection preceded an exacerbation (62). It is also clear that

psychosocial stress could act in concert with immune dysregu-lation to mediate the progression of these disorders as it does inother infectious and autoimmune diseases (6–13).

Our study suggest that a minority of children with TS andearly-onset OCD are sensitive to antecedent GABHS infections,leading to a slight worsening of their tic and OC symptoms.Psychosocial stress continues to be a more potent factor associ-ated with future worsening of tic, OC, and depressive symptoms.In addition to monitoring levels of psychosocial stress, clini-cians should be vigilant to possible infections and other inflamma-tory processes that may compound the impact of psychosocialstressors. Although it is still controversial whether the currentPANDAS criteria can be used to designate a unique clinical entity,we believe that the clinical, epidemiological, and basic scienceobservations support further research into the potential role ofthe innate and adaptive immune systems in the pathogenesis ofa subset of patients presenting with the sudden onset of tics andOCD symptoms (40). In addition, evidence suggests that crucialinteractions exist between central and peripheral dopamine func-tion and the innate and adaptive immune systems (40,63–66).

This research was funded in part by the Tourette SyndromeAssociation (DB), the Donaghue Medical Research Foundation, theYale School of Medicine (JFL), the Echlin Foundation, the Rem-brandt Foundation, Brian Richmand, and the Kaiser Family. Thisresearch was also supported by National Institute of Health GrantNos. MH066187, P01MH049351 (JFL), R01MH061940 (JFL),R01NS42240, MH014235, K05 MH076273 (JFL), K02 MH01527(PJL), DA017713, DA076750, M01RR006022, and RR00125. Theauthors also wish to thank Virginia Eicher, Susan Quatrano, NancyThompson, and Barbara Peterson-Cremer for their invaluableassistance in completing this study.

The authors report no biomedical financial interests or po-tential conflicts of interest.

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