Are gastrointestinal and sleep problems associated with behavioral symptoms of autism spectrum disorder?

Author’s Accepted Manuscript Are Gastrointestinal and Sleep Problems Associated with Behavioral Symptoms of Autism Spectrum Disorder? Xiao-Lei Yang, Shuang Liang, Ming-Yang Zou, Cai-Hong Sun, Pan-Pan Han, Xi-Tao Jiang, Wei Xia, Li-Jie Wu www.elsevier.com/locate/psychres

PII: DOI: Reference:

S0165-1781(17)30092-6 https://doi.org/10.1016/j.psychres.2017.10.040 PSY10940

To appear in: Psychiatry Research Received date: 17 January 2017 Revised date: 18 October 2017 Accepted date: 22 October 2017 Cite this article as: Xiao-Lei Yang, Shuang Liang, Ming-Yang Zou, Cai-Hong Sun, Pan-Pan Han, Xi-Tao Jiang, Wei Xia and Li-Jie Wu, Are Gastrointestinal and Sleep Problems Associated with Behavioral Symptoms of Autism Spectrum Disorder?, Psychiatry Research, https://doi.org/10.1016/j.psychres.2017.10.040 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting galley proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

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Are Gastrointestinal and Sleep Problems Associated with Behavioral Symptoms of Autism Spectrum Disorder? Xiao-Lei Yang, Shuang Liang, Ming-Yang Zou, Cai-Hong Sun, Pan-Pan Han, Xi-Tao Jiang, Wei Xia, Li-Jie Wu* Department of Children’s and Adolescent Health, Public Health College of Harbin Medical University, Harbin 150081, China * Corresponding author: Li-Jie Wu, Department of Children’s and Adolescent Health, Public Health College of Harbin Medical University, 157 Baojian Road, Harbin 150081, China. Tel.: +86-451-87502867

Fax: +86-451-87502867

E-mail: [email protected]

E-mail addresses: [email protected] (Xiao-Lei Yang), [email protected] (Shuang Liang), [email protected] (Ming-Yang Zou), [email protected] (Cai-Hong Sun), [email protected] (Pan-Pan Han), [email protected] (Xi-Tao Jiang), [email protected] (Wei Xia), [email protected] (Li-Jie Wu)

Abstract Many children with autism spectrum disorder (ASD) suffer from concurrent medical symptoms, including gastrointestinal (GI) and sleeping problems. However, there is limited information on the correlation between co-morbidities and autistic behavioral symptoms. In this study, we estimated the prevalence of GI and sleep problems in Chinese ASD children, examined the impacts of GI and sleep problems on autistic behavioral symptoms, and investigated the factors associated with GI and sleep problems. The survey included 169 ASD and 172 healthy children. Data regarding demographic characteristics, GI symptoms, sleep disturbances and behavioral symptoms were collected through questionnaires. GI and sleep problems were prevalent in Chinese ASD children. Moreover, ASD children with GI symptoms reported more severe ASD core symptoms than others. Autistic children’s GI symptoms were associated with maternal sleep problems during pregnancy, child’s 0-6 month food sources and picky eating. ASD children with sleep disturbances had

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lower performance in daily living skills, social cognition, social communication and intellectual development than ASD children without sleep disturbances. Sleep disturbances were associated with extra nutrient supply during lactation and feeding, and child’s picky eating. Autistic children with GI or/and sleep problems may represent clinically relevant subtypes of ASD, for which targeted treatments may be needed.

Keywords: Autism spectrum disorder; Behavior; Gastrointestinal symptoms; Sleep disturbances.

1. Introduction Autism spectrum disorders (ASD) are a complex group of heritable developmental disorders characterized by social deficits, impaired communication, and repetitive stereotypic behavior (Stoner et al., 2014; Young and Barrett, 2015). Autistic symptoms usually emerge during infancy (Jones and Klin, 2013). The prevalence of ASD was 2.24% (1/45) in the United States, according to the 2014 National Health Interview Survey (Zablotsky et al., 2015). Despite a number of studies on the etiology of ASD, the causes in most cases are still unknown (Samsam et al., 2014). Thorough understanding of physiological abnormalities and improving co-morbidities of ASD children will be meaningful to improve the quality of daily life. Moreover, previous studies have reported that ASD children have higher rates of medical co-morbidities than typically developing children (Greenlee et al., 2016; Liu et al., 2016). Aldinger et al. (2015) examined the data from Autism Genetic Resource Exchange and Simons Simplex Collection, and found that ASD children with GI or

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sleep problems had increased risk of other co-morbidities. The high prevalence of GI symptoms in ASD children has been previously documented (Buie et al., 2010) and GI symptoms have been found to be associated with abnormal behavior symptoms (Fulceri et al., 2016) and other psychiatric problems, such as social withdrawal, anxiety (Mazurek et al., 2013), irritability (Greenlee et al., 2016), frustration (Craig and Brown, 1984). Some studies suggested that GI symptoms in ASD may be linked with an imbalance of gut microbiota (Tomova et al., 2015), endocrine stress responses (Ferguson et al., 2016), disruptions in the "gut-brain axis" (Santocchi et al., 2016) and deficiencies in serotonin signaling during development (Marler et al., 2016). Although the high prevalence of GI symptoms in individuals with ASD was frequently reported, the pathogenesis of GI symptoms in ASD children is unclear (Ferguson et al., 2016). Furthermore, there were few studies on the relationship between early life factors of ASD children and GI symptoms. Similarly, sleep disturbances are often problematic for ASD children. A study found autistic children’s sleep disturbances widely effected on daytime life (Malow et al., 2016). Sleep disturbances include insomnia, increased bedtime resistance, sleep disordered breathing, morning awakening and daytime sleepiness problems (Liu et al., 2006). Previous study has revealed the changes in the sleep structure of some ASD children, such as shorter total sleep time and longer sleep latency (Miano et al., 2007). Sleep is an essential part of a healthy lifestyle, especially in developing children, and plays an important role in the early development of the brain (Chen et al., 2015),

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physical growth, immunity, energy recovery, memory consolidation, as well as emotional and cognitive functions and behavioral performance. Thus, it is important to identify which early life factors might influence sleep conditions of ASD children. Although, several studies have reported sleep and/or GI problems in ASD children, there is limited information regarding whether GI and sleep problems are associated with the severity of ASD and behavioral symptoms. The objectives of this study was to: (1) estimate the prevalence of GI and sleep problems in a sample of Chinese ASD children, (2) examine the associations between GI, sleep problems and behavioral symptoms, and (3) investigate risk factors associated with GI and sleep problems in Chinese ASD children.

2. Materials and Methods 2.1. Participants The study was conducted between January 2013 and August 2016 at the Children Development and Behavior Research Center of Harbin Medical University. ASD children who were consecutively visited the Centre were approached and recruited for the study (n = 169) after informed consent. The ASD children were matched with 172 healthy children by sex. All enrolled children were between the ages of 3 and 12. ASD children were diagnosed by a psychiatrist based on the criteria of the Diagnostic and Statistical Manual of Mental Disorders-Fourth Edition (DSM-IV) (American Psychiatric Association, 1994). The exclusion criteria were fragile X syndrome, attention deficit hyperactivity disorder, tic disorder and mental retardation. Control children were recruited from those who attended kindergartens or elementary schools in Harbin, and accepted mental and neurological examinations, which did not find any

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developmental or nervous system diseases. The study was approved by the ethics review committee of Harbin Medical University and written informed consent was obtained from all parents prior to enrollment. 2.2. Measures The ASD and control groups completed clinical information questionnaire and the Children's Sleep Habits Questionnaire (CSHQ). The ASD group also completed the Vineland Adaptive Behavior Scale (VABS), Peabody Picture Vocabulary Test (PPVT), Social Responsiveness Scale (SRS), Chinese version of the Autism Diagnostic Interview–Revised (ADI-R) and Autism Diagnostic Observation Schedule (ADOS), Autism Behavior Checklist (ABC), Childhood Autism Rating Scale (CARS), Pervasive Developmental Disorders Screening Questionnaire (PDDSQ) and Clancy Autism Behavior Scale (CABS). The clinical information questionnaire included child’s early life conditions (child’s food source from birth to 6 month, added solid food time, picky eating habits, medical history and added extra nutrients such as vitamin, calcium and iron etc.), GI conditions (constipation, diarrhea, stool shape, stool smell, flatulence, defecation pattern, abdominal pain), child’s behaviors and psychological development and maternal conditions (sleep problems, diet conditions, picky eating conditions during pregnancy and lactation period and extra nutrients supply including vitamin, calcium and zinc etc.). The CSHQ is used to assess sleep disturbances in children (Johnson et al., 2016)

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and has 33 items with eight dimensions including bedtime resistance, sleep onset delay, sleep duration, sleep anxiety, night waking, parasomnias, sleep disordered breathing and daytime sleepiness (Liu et al., 2015). The total score of above 41 was the optimal clinical cutoff for sleep disturbances. PPVT examines children's receptive vocabulary ability to assess children's intellectual development (Weber et al., 2015). SRS is a brief screening questionnaire for evaluating the severity of social skill deficits and describing other core features of ASD (Frye et al., 2016). SRS has been validated and shown to be reliable and to have good correspondence to the gold-standard ADI-R (Constantino et al., 2003). VABS is used for examining the autistic core behaviors with a focus on communication and adaptive behaviors (Mandic-Maravic et al., 2015). ABC is completed by the parents. It contains five sub-scales (sensory, communication, language, social and self-care skills), which are used to evaluate the severity of autistic symptoms (Krug et al., 1980). CARS is a commonly used tool for diagnosing ASD. The CARS score of 30 to 36 is considered mild to moderate autism, 37-60 is severe autism (Duan et al., 2014). PDDSQ is widely used in China, with excellent reliability and validity. The PDDSQ questionnaire has two versions, one for children 1 to 4 years old and the other for children 4 to 18 years old. CABS was developed in 1969 and introduced to China in the late 1980s. It has been widely used in epidemiological studies in childhood ASD (Sun et al., 2013). 2.3. Statistical analysis The data were analyzed using EpiData 3.02 (EpiData Association, Odense,

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Denmark) and SPSS 17.0 (SPSS Inc., Chicago, IL, USA). For the descriptive data, we computed the means and standard deviations of demographic and outcome variables (independent t test or ANOVA). We used Chi-square, Fisher's exact test and Kruskal-Wallis H test to determine the differences in the distribution of categorical variables (PPVT, CABS, ABC, PDDSQ, CARS, ADIR and ADOS scores) between different subgroups. In order to identify early life factors associated with ASD children’s sleep or GI problems, we computed odds ratio (OR) and 95% confidence intervals (95% CI) by logistic regression analysis. For all analyses, statistical significance was set at P value of 0.05.

3. Results 3.1. Demographic characteristics The ASD group had 169 children (145 males, 24 females, male to female ratio 6.04:1); the mean age was 5.23 ± 2.0 years. The control group included 172 sex matched children (146 males, 26 females, male to female ratio 5.61:1); the mean age was 5.29 ± 1.58 years. 3.2. Gastrointestinal symptoms According to the clinical information questionnaire, 81 (47.93%) among the ASD and 54 (31.39%) among healthy children reported at least one GI symptom, respectively. The prevalence of GI symptoms was significantly higher in the ASD than control groups (OR=2.02; 95% CI: 1.30-3.15, P=0.002). 3.3. GI symptoms and behavioral symptoms in the ASD group In the ASD group, the children with (n = 81) and without (n = 88) GI symptoms

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were significantly different in CARS scores, PDDSQ scores, CABS scores, stereotyped behavior scores of ADI-R, communication and social interaction scores of ADOS (P< 0.05) (Table 1). Table 1 The CARS/PDDSQ/CABS/ADIR/ADOS scores of GI subgroups. GI (n=81)

Non- GI (n=88)

male

65 (80.25)

80 (90.91)

female

16 (19.75)

8 (9.09)

4.99 (1.50)

5.45 (2.36)

30-36

43 (53.09)

63 (71.59)

37-60

19 (23.46)

9 (10.23)

scores-cutoff<10

51 (62.96)

67 (76.14)

scores-cutoff≥10

26 (32.10)

15 (17.05)

<7

19 (23.46)

33 (37.5)

≥7

57 (70.37)

48 (54.54)

≤7

59 (72.84)

76 (86.36)

>7

22 (27.16)

12 (13.64)

scores-cutoff<5

67 (82.72)

82 (93.18)

scores-cutoff≥5

14 (17.28)

6 (6.82)

scores-cutoff<5

63 (77.78)

79 (89.77)

scores-cutoff≥5

18 (22.22)

9 (10.23)

Item

2/t

P

3.94

0.047

0.13

0.131

6.64

0.010

4.97

0.026

4.39

0.036

4.80

0.028

4.43

0.035

4.52

0.033

Gender, n (%)

Age (years), mean (SD) CARS scores, n (%)

PDDSQ scores, n (%)

CABS scores, n (%)

ADI-R scores, n (%) stereotyped behavior

ADOS scores, n (%) communication

social interaction

Note: CARS was available for 162 (95.86%), and twenty-eight ASD children’s CARS scores were less than 30; PDDSQ was available for 159 (94.08%); CABS was available for 157 (92.90%).

3.4. Sleep disturbances in two groups Total CSHQ scores over 41 were considered the optimal clinical cutoff for sleep disturbances. According to CSHQ total scores for the two groups, there were 152

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(89.94%) and 124 (72.09%) children with sleep disturbances in the ASD and control groups respectively; the prevalence of sleep disturbances was significantly higher in the ASD group than that in the control group (OR=3.46; 95% CI: 1.90-6.31, P<0.001). By examining specific sleep disturbances, ASD children were more likely to report bedtime resistance, sleep onset delay, sleep anxiety, night waking, parasomnias and sleep disordered breathing than control children (P<0.05) (Table 2) Table 2 The scores of CSHQ in the ASD and control groups. ASD (n=169)

Control (n=172)

Mean (SD)

Mean (SD)

Bedtime resistance

11.40 (2.52)

10.12 (2.56)

<0.001

Sleep onset delay

1.75 (0.76)

01.59 (0.67)

0.043

Sleep duration

4.12 (1.38)

03.95 (1.43)

0.262

Sleep anxiety

7.10 (1.63)

06.67 (1.87)

0.026

Night waking

4.20 (1.32)

03.73 (0.97)

<0.001

Parasomnias

8.53 (1.51)

08.11 (1.25)

0.006

Sleep disordered breathing

3.74 (1.06)

03.51 (0.81)

0.022

Daytime sleepiness

12.46 (2.50)

12.36 (2.60)

0.731

CSHQ total scores

48.92 (5.79)

46.79 (6.12)

0.001

Item

P

3.5. Sleep disturbances and behavioral symptoms in the ASD group Among ASD children, the data showed that children with (152; 89.94%) and without (17; 10.06%) sleep disturbances significantly differed in PPVT scores, VABS total standard scores, daily living skills and socialization item scores of VABS, ABC scores, PDDSQ scores, SRS total scores, social cognition and social communication item scores of SRS (P< 0.05) (Table 3) Table 3 The VABS/SRS /PPVT/ABC/PDDSQ scores of sleep subgroups. Sleep (n=152)

Non- Sleep (n=17)

male

130 (85.53)

15 (88.24)

female

22 (14.47)

2 (11.76)

Item

2 /t/H/ Fisher test

P

Children gender, n (%) -

1.00

10

5.15 (1.91)

5.93 (2.64)

1.53

0.127

total standard scores

72.13 (19.77)

89.41 (25.99)

3.31

0.001

daily living skills

73.64 (20.13)

86.12 (24.77)

2.37

0.019

socialization

63.83 (16.70)

77.47 (27.49)

2.96

0.004

total scores

86.13 (24.32)

72.53(16.32)

2.25

0.026

social cognition

18.46 (4.50)

15.94 (3.47)

2.02

0.045

social communication

32.42 (8.87)

26.29 (8.82)

2.70

0.008

<70

119 (78.29)

5 (29.41)

70-85

22 (14.47)

7 (41.18)

-4.38

<0.001

>85

11 (7.24)

5 (29.41)

≤53

36 (23.68 )

8 (47.06)

54-66

22 (14.47)

6 (35.29)

-3.09

0.002

≥67

88 (57.89)

3 (17.65)

scores-cutoff<10

101 (66.45)

17 (100)

scores-cutoff≥10

41 (26.97)

0

-

0.007

Age (years), mean (SD) VABS scores, mean (SD)

SRS scores, mean (SD)

PPVT scores, n (%)

ABC scores, n (%)

PDDSQ scores, n (%)

Note: ABC was available for 163 (96.45%); PDDSQ was available for 159 (94.08%).

3.6. The prevalence of both co-morbidities in the ASD and control groups Among the ASD group, 73 children (43.20%) reported both GI and sleep co-morbidities, which is significantly higher than that in the control group (23.84%). Statistical analysis found that the proportions of both co-morbidities, only GI symptoms, only sleep disturbances, and non-GI and non-sleep problems were statistically different between the ASD and control groups (P<0.001) (Table 4). Table 4 The co-morbidity conditions in the ASD and control groups.

ASD N (%)

Control N (%)

Both co-morbidities

73(43.20)

41(23.84)

Only GI symptoms

8(4.73)

13(7.56)

79(46.75)

83(48.25)

9(5.32)

35(20.35)

Item

Only sleep disturbances Non-GI and non-sleep problems

P

<0.001

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3.7. Both GI and sleep co-morbidities and behavioral symptoms in the ASD group We examined the relationships between GI/sleep problems and behavioral symptoms in the ASD group. Participants in the ASD group were divided into both co-morbidities, only GI symptoms, only sleep disturbances and non-GI and non-sleep problems subgroups. Through this subdivision, it was found that PPVT scores, VABS total standard scores, socialization scores of VABS, ABC scores, CARS scores, PDDSQ scores, CABS scores, communication and social interaction of ADOS scores were statistically different in the four subgroups (P<0.05). Through multiple comparisons between different subgroups, we found that PPVT scores were statistically different between both co-morbidities and non-comorbidity subgroups (P=0.012). Specifically, the ABC scores of both co-morbidities and only sleep subgroups were significantly different from that of non-comorbidity subgroup (P=0.009&0.002) (Table 5). Table 5 The VABS/PPVT/ABC/CARS/PDDSQ/CABS/ADOS scores of four subgroups. Both

Item

co-morbidities (n=73)

Only GI

Only sleep

(n=8)

(n=79)

Non-GI and non-sleep (n=9)

2/t

/H

P

Children gender, n (%) male

59(80.82)

6(75)

71(89.87)

9(100)

female

14(19.18)

2(25)

8(10.13)

0

4.98(1.53)

5.07(1.23)

5.31(2.20)

total standard scores

74.05 (21.16)

95.25(27.90)

socialization

65.25(18.77)

<70 70-85

Age (years), mean(SD)

4.51

0.157

6.69(3.35)

2.10

0.103

70.34(18.36)

84.11(24.31)

4.47

0.005

90(33.79)

62.52(14.52)

66.33(14.68)

5.87

0.001

58(79.45)a#

1(12.5)

61(77.22)

4(44.44)

11(15.07)

5(62.5)

11(13.92)

2(22.22)

20.81

<0.001

4(5.48)

2(25)

7(8.86)

3(33.34)

18(24.66)a#

4(50)

18(22.78)b#

4(44.44)

11.42

0.010

VABS scores, mean(SD)

PPVT scores, n (%)

>85 ABC scores, n (%) ≤53

12

54-66

14(19.18)

1(12.5)

8(10.13)

5(55.56)

≥67

39(53.42)

3(37.5)

49(62.03)

0

30-36

40(54.79)

3(37.5)

56(70.89)

7(77.78)

37-60

19(26.03)

0

9(11.39)

0

scores-cutoff <10

43(58.90)

8(100)

58(73.42)

9(100)

scores-cutoff ≥10

26(35.62)

0

15(18.99)

0

<7

15(20.55)

4(50)

28(35.44)

5(55.56)

≥7

54(73.97)

3(37.5)

44(55.70)

4(44.44)

scores-cutoff <5

59(80.82)

8(100)

75(94.94)

7(77.78)

scores-cutoff ≥5

14(19.18)

0

4(5.06)

2(22.22)

scores-cutoff < 5

55(75.34)

8(100)

70(88.61)

9(100)

scores-cutoff ≥5

18(24.66)

0

9(11.39)

0

CARS scores, n (%) 7.79

0.035

11.36

0.007

9.15

0.022

8.94

0.020

7.02

0.048

PDDSQ scores, n (%)

CABS scores, n (%)

ADOS scores, n (%) communication

social interaction

Note: #p＜0.0167, a. Both co-morbidities subgroup compared with non-GI and non-sleep problems subgroup, b. Only sleep subgroup compared with non-GI and non-sleep problems subgroup. ABC was available for 163 (96.45%); CARS was available for 162 (95.86%), and twenty-eight ASD children’s CARS scores were less than 30; PDDSQ was available for 159 (94.08%); CABS was available for 157 (92.90%).

3.8. Multivariate analysis of GI symptoms in the ASD group We examined the factors associated with the GI symptoms in the ASD group. Logistic regression analysis showed that gender (OR=3.88, 95% CI: 1.33–11.35), maternal sleep problems during pregnancy (OR=4.45, 95% CI: 1.72-11.56), child supplied infant formula and breastfeeding from birth to 6 month (OR=3.17, 95% CI: 1.20-8.36), child supplied only infant formula from birth to 6 month (OR=5.04, 95% CI: 1.43-17.78) and child’s picky eating (OR=2.74, 95% CI: 1.28-5.88) were significantly associated with GI symptoms in the ASD group (Table 6). Table 6 Multivariate analysis for participants with and without GI symptoms. Variable

Wald 2

OR

95% CI

P

13

Gender male female

6.14

3.88

1.33-11.35

0.013

9.42

4.45

1.72-11.56

0.002

3.01

2.05

0.91-4.59

0.083

infant formula + breastfeeding

5.43

3.17

1.20-8.36

0.020

infant formula

6.31

5.04

1.43-17.78

0.012

2.91

1.99

0.90-4.39

0.088

6.68

2.74

1.28-5.88

0.011

1.07

1.66

0.64-4.31

0.301

Sleep problems during pregnancy no yes Maternal picky eating during lactation no yes Child’s 0-6 month food sources breastfeeding

Add solid food time ≥6 month <6 month Child’s picky eating no yes Child’s allergy history no yes

3.9. Multivariate analysis of sleep disturbances in the ASD group Logistic regression analysis demonstrated that extra nutrients supply for mother during lactation (OR=0.30, 95%CI: 0.10-0.93), extra nutrients supply for child during feeding (OR=0.24, 95%CI: 0.06-0.91) and child’s picky eating (OR=4.00, 95%CI: 1.16-13.80) were associated with sleep disturbances in the ASD group (Table 7). Table 7 Multivariate analysis for participants with and without sleep disturbances. Variable

Wald 2

OR

95% CI

P

0.52

2.21

0.26-19.10

0.472

Gender male female Extra nutrients for mother during lactation no

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yes

4.32

0.30

0.10-0.93

0.038

4.39

0.24

0.06-0.91

0.036

4.82

4.00

1.16-13.80

0.028

Extra nutrients for child during feeding no yes Child’s picky eating no yes

4. Discussion To the best of our knowledge, this is the first study on GI and sleep co-morbidities in Chinese ASD children. We found that ASD children were more likely to have GI symptoms and sleep disturbances than the control group. We observed a higher percentage of ASD (47.93%) versus control (31.39%) group who reported with GI symptoms (P=0.002), which was consistent with the result of the previous study (Fulceri et al., 2016). Many autistic children are nonverbal or minimally verbal and may not communicate information about their GI symptoms so clearly as healthy children (Buie et al., 2010). Of course, ASD children often have psychological and affective problems, which are associated with GI symptoms (Mazefsky et al., 2014; Mazurek et al., 2013). Valicenti-McDermott et al. (2006) found that 70% ASD children had GI symptoms which is higher than that of our research (47.93%). However, a recent study showed that the prevalence of GI symptoms in autistic children ranged from 23% to 70% (Chaidez et al., 2014). The heterogeneity in prevalence may be due to the differences in clinical criteria, study design and population characteristics. Of note, our research used a large sample, not only provided a better understanding of the prevalence of GI symptoms in Chinese ASD children but also evaluated their behavioral characteristics. We found that GI

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symptoms correlated with the severity of autistic behaviors by the results of assessment scales, which is consistent with what Samsam et al. (2014) and Peters et al. (2014) reported. However, one study didn’t find the connection between ASD children’s GI problems and autistic symptom severity among 95 children with high-functioning autism (Mazefsky et al., 2014). One study showed that the autistic children with GI dysfunction exhibited language and social impairments (Gorrindo et al., 2012), but we did not find the same result about language difference between GI and non-GI subgroups in ASD children. Nevertheless, we found that social interaction, communication and stereotyped behavior problems in the GI subgroup were more serious than that of the non-GI subgroup. It is worth noting that this study not only investigated the GI symptoms, but also focused on their sleep disturbances. Our survey found that the prevalence of sleep disturbances (89.94%) in Chinese ASD children was higher than that of other research. Liu et al. (2006) and Krakowiak et al. (2008) reported that approximately 86% and 53% of ASD children, respectively, have at least one sleep problem. The discrepancies may be due to different questionnaires, populations and response times. In China, children often sleep with their parents who commonly pay attention on the sleeping progress of their children, which may contribute to the higher detection rate of sleep disturbances in Chinese children than that reported elsewhere. We also found that bedtime resistance, sleep onset delay, sleep anxiety, night waking, parasomnias, sleep disordered breathing and CSHQ total scores were significantly different between the ASD and control groups (P<0.05). In addition, the mean dimensional scores of the

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CSHQ questionnaire in the ASD group were higher than in the control group. Sleep disturbances may affect brain development, and even lead to abnormal behaviors that adversely influence normal life and learning. Our results showed that ASD children with sleep disturbances had serious autistic behaviors, especially in the domains of daily living skills, social cognition, social communication and intelligence quotient. Sleep disturbances could aggravate autistic behaviors, such as social skill deficits, communication impairments (Levin and Scher, 2016) and stereotypic behaviors (Cohen et al., 2014). May et al. (2015) found that reduced sleep disturbances were associated with improved social ability. Although the correlation between sleep and autistic behaviors has been identified, more evidence is needed to explore the causality. Based on the aforementioned results, we further analyzed our dataset, finding the prevalence of both sleep and GI co-morbidities was 43.20% in the ASD group, while only 23.84% in the control group. The proportions of the both co-morbidities, only GI symptoms, only sleep disturbances and without GI or sleep problems subgroups were remarkably different between the ASD and control groups (P<0.001). In addition, the data suggested that co-occurrence of sleep and GI problems may have a greater impact on ASD children's intellectual development. Although a number of studies suggest the links between neurologic impairments and GI symptoms or sleep disturbances, these relationships remain unproven (Coury et al., 2009). Our research indicated that both co-morbidities and only sleep subgroups had more severe autistic behaviors than non-co-morbidities subgroup. Despite the direction of the association

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between both co-morbidities and ASD core symptoms is unclear, it is possible that effective treatment for either of co-morbidities or autistic core symptoms might have beneficial effects on the other one. Multivariate analysis was used to identify early factors that are associated with sleep and GI problems in the ASD group. It was found that females were more likely to have GI symptoms, which was consistent with the results of Cilleruelo et al. (2016). Additionally, we found that if the mother had sleep problems (sleep time decreased, difficulty falling asleep, night waking etc.) during pregnancy, her child was more likely to have GI symptoms. GI symptoms were also more likely to occur in the children who fed on only infant formula or breastfeeding and infant formula during first 6 months from birth. By logistic regression analysis, child’s picky eating was associated with GI symptoms in the ASD group. Numerous studies have reported that ASD children are more likely to have picky eating habits (Bandini et al., 2010; Steward-Scott, 2014), which leads to inadequate nutrient intake. Interestingly, our result revealed that child’s picky eating was related to both GI and sleep problems. Additionally, extra nutrients supply for ASD child’s mother during lactation, her child was not prone to sleep disturbances. ASD children who were given extra nutrient during the period of being fed were less likely to have sleep disturbances than others. Bandini et al. (2010) noted that nutrient deficiency of autistic children were common, such as vitamin D, vitamin E, and calcium. The recent review showed that vitamin D plays the role of a sleep modulator (de Oliveira et al., 2017). A large study on the level of vitamin D in adolescents found that poor quality sleep was associated with

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vitamin D deficiency (Ataie-Jafari et al., 2015). Based on evidence above, identification of factors associated with GI and sleep problems in ASD children might be beneficial to improve both conditions. Above all, the identification of physiological abnormalities in ASD could lead to a better understanding of the biological dysfunctions which affect behavioral symptoms, and formulate individualized treatment plans (Frye and Rossignol, 2016). Although none of the co-morbidities were specific to ASD, our findings could explain, at least in part, the GI and sleep co-morbidities are associated with ASD behavioral symptoms. Consequently, autistic children with GI and/or sleep co-morbidities might be regarded as clinical subtypes of ASD. Treatment of both co-morbidities could contribute to the prognosis of ASD. This study had several limitations. First, this study was cross-sectional. Thus causal inferences cannot be made. Second, the ASD group consisted of children who came to our center for medical diagnoses or behavioral interventions. One should be cautious regarding the generalization of the results. Furthermore, the small sample for ASD children who have no sleep disturbances might affect the results. Third, parts of the questionnaires were completed by caregivers, who had different educational levels and attitudes, which resulted in several questionnaires not meeting the criteria for inclusion. Finally, customs, family life styles and the seasons of investigation might also indirectly affect the results of the study. 5. Conclusion Three important implications can be drawn from our results. First, both GI and

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sleep co-morbidities were prevalent in Chinese ASD children. Second, autistic children with GI and/or sleep problems presented more serious behavioral symptoms, and ASD children with sleep disturbances also had poor intellectual development. Finally, through multivariate analysis, we found that ASD children’s GI symptoms were associated with maternal sleep problems during pregnancy, child’s picky eating, infant formula only and breastfeeding mixed with infant formula during first 6 months from birth. Meanwhile, ASD children’s sleep disturbances were associated with extra nutrient supply during lactation and feeding, and child’s picky eating. In conclusion, these findings contribute to our understanding of the associations between GI and/or sleep co-morbidities and the severity of ASD behaviors. Moreover, our results may contribute to ASD treatments and rehabilitation.

Acknowledgments The participation of the families is gratefully acknowledged. The authors extend thanks to the Children Development and Behavior Research Center of Harbin Medical University for their cooperation. This work was supported by National Basic Research Program (973 Program) (Nos. 2012CB517901).

Conflict of Interest The authors declare no conflict of interest.

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Highlights  Gastrointestinal and sleep problems are prevalent in Chinese children with autism spectrum disorder. 

Autistic children with gastrointestinal and/or sleep problems presented more serious behavioral symptoms.

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

Autistic children’s gastrointestinal symptoms were associated with maternal sleep problems during pregnancy, child’s 0-6 month food sources and child’s picky eating.

 ASD children’s sleep disturbances were associated with extra nutrient supply during lactation and feeding, and child’s picky eating.  Autistic children with gastrointestinal or/and sleep problems may represent clinically relevant subtypes of ASD, for which targeted treatments may be needed.