Predictors of recurrence of major depressive disorder.

A total of 201 patients with major depressive disorder from four hospitals in Malaysia were followed up for 5 years to determine the prognostic factors of recurrent major depressive disorder that could potentially contribute to improving the management of MDD patients. For each individual patient, at the time of recruitment as part of a case-control study, information was collected on recent threatening life events, personality and social and occupational functioning, while blood samples were collected to genotype single nucleotide polymorphisms of vitamin D receptor (VDR), zinc transporter-3 (ZnT3), dopamine transporter-1 (DAT1), brain-derived neurotropic factor (BDNF), serotonin receptor 1A (HT1A) and 2A (HT2A) genes. Kaplan-Meier and Cox-regression were used to estimate hazard functions for recurrence of major depressive disorder. Individuals with severe MDD in previous major depressive episodes had five and a half times higher hazard of developing recurrence compared to mild and moderate MDD (HR = 5.565, 95% CI = 1.631-18.994, p = 0.006). Individuals who scored higher on social avoidance had three and a half times higher hazard of recurrence of MDD (HR = 3.525, 95% CI = 1.349-9.209; p = 0.010). There was significant interaction between ApaI +64978C>A single nucleotide polymorphism and severity. The hazard ratio increased by 6.4 times from mild and moderate to severe MDD for A/A genotype while that for C/A genotype increased by 11.3 times. Social avoidance and severity of depression at first episode were prognostic of recurrence. Screening for personality factors at first encounter with MDD patients needs to be considered as part of the clinical practice. For those at risk of recurrence in relation to social avoidance, the psychological intervention prescribed should be customized to focus on this modifiable factor. Prompt and appropriate management of severe MDD is recommended to reduce risk of recurrence.

Introduction

Major depressive disorder (MDD) is defined as “discrete episodes of at least 2 weeks’ duration with clear-cut changes in affect, cognition, and neuro-vegetative functions, and inter-episode remissions” and often is recurrent in nature [1]. Recurrence is the return of symptoms after at least 2 consecutive months between separate episodes during which time criteria are not met for a major depressive episode (MDE) and there must be the return of at least 5 out of 9 symptoms of depression [1]. Many patients are at substantial risk of later recurrence, with 60% lifetime risk of recurrence after the first major depressive episode. As many as 70% of those with 2 MDEs have recurrences throughout their life, and 90% of those with three or more episodes will experience further recurrent episodes [2]. In addition, one third to half of the patients had recurrence within one year of discontinuation of treatment [3]. Each recurrence also carries a 10–20% risk of becoming unremitting and chronic [4]. Recurrent MDD in turn increases risk of significant functional impairment, suicide and comorbid physical health problems [5-9], incurring heavy health and economic burdens [10].

Many studies showed that sociodemographic factors such as younger age, women, and those who have never been married were at higher risk of recurrence [7, 11–15]; however, results of other studies on these risk factors have been inconsistent. A systematic review by Hardeveld and colleagues (2009) did not find any association between sociodemographic factors such as age, gender, socioeconomic status, marital status and risk of recurrence [16, 17, 18]. However, clinical variables such as number of previous episodes [7, 11, 19–22], severity of last episode [21, 22] and family history of MDD [21– 23] were strong predictors of recurrence. In addition, patients who had residual depression symptoms despite responding to treatment, had higher risk of recurrence (Serafini et al., 2018; Nierenberg et al., 2010; Judd et al., 1998) [19, 24, 25]. Studies also showed that personality traits were related to risk of recurrence [26]; Nobbelin et al. (2018) reported that patients with premorbid nervous or tense personalities had greater likelihood of recurrence [27] (Nobbelin et al., 2018).

Burcasa and Iacono in their review suggested that vulnerability to recurrent MDD has a genetic component [28]. Mutations in the brain-derived neurotropic factor (BDNF) [29, 30] and serotonin receptor genes [31] were shown to be associated with recurrent MDD. In addition, findings by Kuningas et al. (2009) [32] and Lye et al. [33] that single nucleotide polymorphisms (SNPs) of vitamin D receptor (VDR) and zinc transporter-3 (SLC30A3) genes were respectively associated with MDD, and together with evidence that SNPs of dopamine transporter-1 gene increased risk of MDD[34, 35], led to renewed interest in determining if an association exists between VDR, ZnT3 SNPs and recurrence of MDD.

Although acute management of MDDs is generally effective, many patients experience recurrence following remission. Identifying prognostic factors in patients at high risk of recurrence provides a window of opportunity for specific secondary prevention as well as initiation of long-term maintenance treatment. The objective of the present study is to determine the prognostic factors of recurrence that could potentially contribute to improving the management of patients with MDD.

Materials and methods

Ethics statement

This study was approved by the Medical Research Ethics Committee of the Ministry of Health Malaysia (NMRR No.: NMRR-14-688-19696). Written informed consent was obtained from all the participants.

Data collection

The original study was a case-control study in which the methodology has been described elsewhere [33]. Inclusion criteria included male and female patients from 18 to 65 years of age, who were diagnosed with single or recurrent, non-psychotic episode of Major Depressive Disorder (using the Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition (DSM-5) [1]) diagnosed less than 2 years prior to recruitment. Patients who had significant suicidal risk as assessed by the psychiatrist or diagnosed with dementia, schizophrenia or other psychotic disorder, bipolar I or II disorder, or anxiety disorders including panic disorder, generalized anxiety disorder, obsessive-compulsive disorder and post-traumatic stress disorder, were excluded from the study.

Baseline sociodemographic characteristics as well as comorbidities (such as hypertension, diabetes mellitus, heart disease, stroke, cancer and asthma) were captured. Standardized questionnaires such as Lists of Threatening Events (LTEs) [34] and Temperament and Personality (T&P) [36] were used. The English version of the 12-item LTE questionnaire was reported to have good internal consistency and test-retest reliability (Cronbach's alpha = 0.84; Cohen's kappa = 0.72), with sensitivity of 89% and specificity of 74% [37]; the Malay version by Ng et al. (2009) was shown to be reliable as well (Kappa ĸ = 0.7 to 0.9) [38].

The T&P questionnaire consisted of 109 questions to capture information on eight personality dimensions and two personality functioning dimensions with Cronbach's α coefficients ranging from 0.62 to 0.91 and intra-class correlations from 0.72 to 0.93 [36, 39]. Items measuring each dimension add up to a scaled score, in which different cut off values were used to determine the tendencies of the dimensions. Cut off points for the different dimensions were as follows: Anxious worrying (18 and above—indicates a greater tendency to become stressed, worried and anxious); Personal reserve (17 and above—is associated with a tendency to keep one’s inner feelings to oneself); Perfectionism (31 and above—is associated with a tendency to be very responsible, to have high standards for oneself and to be highly committed to tasks and duties); Irritability (21 and above—is associated with a tendency to be quick-tempered and to externalise stress by becoming snappy and irritated by little things); Social avoidance (17 and above—is associated with a tendency to be introverted); Interpersonal sensitivity (14 and above—is associated with a tendency to worry about rejection or abandonment); Self-criticism (10 and above—is associated with a tendency to be very tough on oneself); Self-focus (9 and over—indicating prioritizing one’s own needs over other peoples’); Cooperativeness (20 and above—is associated with a tendency to be generally helpful); and Effectiveness (18 and above—indicates an ability to cope well with different situations and to be confident in problem solving). Higher scores indicate a higher tendency for those dimensions. A clinical record form endorsed by a psychiatrist (NI) (S1 Appendix) was used to identify recurrence based on the definition mentioned above and the symptoms reported at each visit, time from first diagnosis to occurrence of first episode of recurrence of MDD, duration of first episode of recurrence, subsequent episodes of MDD following the first episode of recurrence, severity of first and subsequent MDDs, full or partial remission for each recurrent episode, and family history of MDE. Severity was classified as mild, moderate and severe, according to the number symptoms based on ICD-10 [40], taking into consideration the level of severity of those symptoms and the degree of functional disability using DSM-5 [1]. Mild MDD was defined as presence of two or three MDD symptoms, which were distressing but manageable with minor functional impairment; moderate MDD was characterized by four or more MDD symptoms and patients were functionally impaired; severe MDD was defined by several marked and distressing MDD symptoms which were seriously distressing and unmanageable, with severe functional impairment associated with loss of self-esteem and ideas of worthlessness or guilt, suicidal thoughts or acts [1]. Full remission is defined by DSM-5 as having no significant signs or symptoms of MDD in the past 2 months [1]. Partial remission is defined as presence of symptoms of the immediate previous MDE but they do not fulfil full criteria for MDD or “there is a period lasting less than 2 months without any significant symptoms of MDE following the end of such an episode” [1].

Blood collection

5 mL of blood was collected using evacuated ethylenediamine tetra acetic acid (EDTA) tubes (Vacutainer Tubes, Becton-Dickinson, NJ, USA) from each subject, was stored under 4°C and processed on the same day.

Extraction of genomic DNA

We used the QIAamp DNA Mini Kit (QIAgen, USA) to extract genomic DNA [41]. 25μL of proteinase K and 200μL of lysis buffer were added to the buffy coat and incubated at 56°C for 30 minutes to maximize the cell lysis. We then precipitated DNA in 200μL of absolute ethanol before spinning through a filtered spin column. The column was washed twice in AW1 and AW2 wash buffer, followed by a drying spin at maximum speed for 1 minute. DNA was eluted with 50μL of nuclease free water and DNA yield. We then purified and quantified the extracted genomic DNA using a NanoPhotometer®Classic (Implen, USA). DNA purity of all samples were assessed using the 260/280nm ratio within the range of 1.7–2.0.

Determination of Vitamin D Receptor, zinc transporter-3, Dopamine Transporter-1, Brain-derived neurotropic factor, serotonin receptor 1A and 2A Single Nucleotide Polymorphisms (SNPs)

Primers (both sense and antisense) were used to amplify the targeted single nucleotide polymorphisms (SNPs) namely, BsmI +63980 G>A (rs1544410), ApaI +64978C>A (rs7975232) and TaqI +65058 T>C (rs731236) in the Vitamin D Receptor (VDR) gene, SLC30A3 rs11126396 of ZnT3 gene, rs40184 C>T of Dopamine Transporter-1 (DAT1) gene, rs6265 G196A of Brain-derived neurotropic factor (BDNF) gene, rs6295 -1019C>G of serotonin receptor 1A(HT1A) gene, and rs6311 -1438A>G of serotonin receptor 2A (HT2A) gene. Details of the genotyping methods have been fully described elsewhere [33, 42] and in S2 Appendix and S3 Appendix.

Data analysis

Relative frequencies were used to describe variables studied including sociodemographic factors and distribution of single nucleotide polymorphisms of all the genes. Chi-square test or Fisher’s exact test were used to determine differences in sociodemographic variables between recurrent patients and non-recurrent patients. Kaplan-Meier analysis was used to perform univariate analysis for sociodemographic variables, family history of psychiatric illnesses, severity of first MDE, comorbidities, and SNPs of BsmI (rs1544410), TaqI (rs731236) and ApaI (rs7975232) of VDR gene, SLC30A3 rs11126396 of ZnT3 gene, rs40184 of DAT1 gene, rs6265 of BDNF gene, rs6295 of HT1A, rs6311 HT2A gene, LTE and T&P domains. Subject(s) who did not experience recurrent MDE at the end of the study period were censored. From the Kaplan-Meier results, variables with p values less than 0.25 were then selected for the Cox regression model which estimated hazard ratios [43, 44, 45].

The Cox-proportionate hazards model was used in regressing time to recurrence on severity of first MDE, social avoidance, irritability, anxious worrying, interpersonal sensitivity, self-criticism, effectiveness, age of first MDE, gender, educational level, family history of psychiatric illness, and comorbidities. Interactions between the significant predictor variables and genotypes of single nucleotide polymorphisms of VDR genes namely, BsmI +63980 G>A, ApaI +64978C>A and TaqI +65058 T>C, and SLC30A3 rs11126396 of ZnT3 gene, rs40184 of DAT1 gene, rs6265 of BDNF gene, rs6295 of HT1A and rs6311 HT2A gene were tested. Statistical significance was set at alpha of 0.05.

Results

Over the five-year period, of the 201 patients traced from the case histories, 145 patients were available for survival analysis. 20.9% of the MDD patients suffered first recurrence; the rest were censored. Table 1 shows that there was no difference between the patients with data available for survival analysis and those for whom records were not available. Table 2 depicts the baseline information among those with and without recurrence. The age distribution was similar among both groups. The proportion of patients who experienced recurrence was slightly higher in females compared to males, with a female to male ratio of 1.5:1. Recurrence of MDD was also higher in those with academic degrees and postgraduate qualifications. However, those differences were not statistically significant.

Table 1

Comparison of cases included for analysis versus missing cases.

Variables	Followed up		Missing		Statistica	p- value
	n	%	n	%
Race					1.354	0.508
Malay	100	51.0	49	47.1
Chinese	60	30.6	30	28.8
Indian and others	36	18.4	25	24.0
Gender					0.765	0.382
Male	60	30.6	37	35.6
Female	136	69.4	67	64.4
Education level					0.638	0.888
Primary or	95	48.5	52	50.0
Secondary
Certificate	18	9.2	7	6.7
Diploma	30	15.3	15	14.4
Degree/Postgraduate	53	27.0	30	28.8
Comorbidities					0.004	0.949
Yes	69	35.2	37	35.6
No	127	64.8	67	64.4
Family history					0.918	0.632
Yes	54	27.6	26	25.0
No	124	63.2	71	68.3
Unsure	18	9.2	7	6.7
TaqI					0.693	0.707
T/T	139	70.9	78	75.0
T/C	44	22.4	21	20.2
C/C	13	6.6	5	4.8
ApaI					0.123	0.940
A/A	41	20.9	20	19.2
A/C	75	38.3	41	39.4
C/C	80	40.8	43	41.4
Anxious worrying					0.75	0.386
High tendency	51	26.6	21	21.9
Low tendency	141	73.4	75	78.1
Irritability					2.963	0.085
High tendency	25	13.0	20	20.8
Low tendency	167	87.0	76	79.2
Social avoidance					1.169	0.280
High tendency	16	8.4	12	12.4
Low tendency	175	91.6	85	87.6
Interpersonal					0.097	0.755
sensitivity	57	30.5	31	32.3
High tendency	130	69.5	65	67.7
Low tendency
Self-criticism					0.629	0.428
High tendency	140	72.9	65	68.4
Low tendency	52	27.1	30	31.6
Effectiveness					0.129	0.719
High tendency	60	33.3	29	31.2
Low tendency	120	66.7	64	68.8
Mean Age, years (sd)	196	38.9 (13.01)	104	38.5 (11.93)	0.228*	0.820

a The statistic reported for all variables was the chi-square value, except the variable labelled * (t-statistic)

Table 2

Sociodemographic characteristics of study population by recurrence (n = 145).

Variables	Recurrencen (%)		No Recurrence n (%)		χ 2	p value
Age (years old)					0.017	0.898
18–39	24	21.2	89	78.8
40–65	17	20.5	66	79.5
Race					1.051	0.305
Malay	18	18.0	82	82.0
Others	23	24.0	73	76.0
Gender					1.831	0.176
Male	9	15.0	51	85.0
Female	32	23.5	104	76.5
Education level					1.672	0.196
Primary /Secondary/Certificate	20	17.7	93	82.3
Diploma/Degree/Postgraduate	21	25.3	62	74.7

Severity, social avoidance and interaction between ApaI polymorphism and severity significantly prognosticated recurrence (Table 3). Individuals with severe MDD at the first MDE had five and a half times higher hazard of developing recurrence compared to mild and moderate MDD (HR = 5.565, 95% CI = 1.631–18.994, p = 0.006). Individuals with a higher score of social avoidance had three and a half times higher hazard of recurrence of MDD (HR = 3.525, 95% CI = 1.349–9.209; p = 0.010). The hazard functions for severity and social avoidance are depicted in Fig 1 and Fig 2 respectively.

Table 3

Cox proportionate hazard analysis for predictors of MDD recurrence (n = 145).

Variables		Recurrencen (%)		No Recurrence n (%)		Adjusted HRa	95% CI	p value
Age group	18–39	24	21.2	89	78.8	1
	40–65	17	20.5	66	79.5	0.778	(0.323–1.874)	0.575
Gender	Male	9	15.0	51	85.0	1
	Female	32	23.5	104	76.5	0.996	(0.401–2.476)	0.994
Educational level	Primary / Secondary/Certificate	20	17.7	93	82.3	1
	Diploma/Degree/ Postgraduate	21	25.3	62	74.7	1.252	(0.544–2.883)	0.597
Comorbidities	Yes	10	14.3	60	85.7	1
	No	31	23.7	100	76.3	0.936	(0.376–2.334)	0.888
Family History of psychiatric illness	Yes	45	83.3	9	16.7	1
	No	102	79.1	27	20.9	1.937	(0.804–4.666)	0.141
	Unsure	13	72.2	5	27.8	1.756	(0.478–6.448)	0.397
Severity	Mild to Moderate	20	17.4	95	82.6	1
	Severe	21	25.9	60	74.1	5.565	(1.631–18.994)	0.006*
Anxious Worrying	Low	27	19.0	115	81.0	1
	High	13	26.0	37	74.0	0.769	(0.285–2.071)	0.603
Irritability	Low	32	19.2	135	80.8	1
	High	7	28.0	18	72.0	1.328	(0.413–4.274)	0.634
Social Avoidance	Low	31	17.7	144	82.3	1
	High	8	50.0	8	50.0	3.525	(1.349–9.209)	0.010*
Interpersonal Sensitivity	Low	27	20.6	104	79.4	1
	High	12	21.4	44	78.6	0.942	(0.371–2.394)	0.900
Self-criticism	Low	4	7.5	49	92.5	1
	High	36	25.9	103	74.1	2.964	(0.810–10.850)	0.101
Effectiveness	Low	27	22.5	93	77.5	1
	High	10	16.7	50	83.3	1.048	(0.435–2.523)	0.917
TaqI	T/T	33	23.9	105	76.1	1
	T/C	7	15.6	38	84.4	0.700	(0.258–1.900)	0.483
	C/C	1	7.7	12	92.3	0.141	(0.016–1.264)	0.080
ApaI*Severity at first MDE
	ApaI*Severity					1
	b ApaI (1)*Severity					0.582	(0.181–1.876)	0.365
	b ApaI (2)*Severity					0.122	(0.024–0.610)	0.010*

aHazard ratio, controlling for age and gender

b Dummy variables: ApaI (1) = C/A; ApaI (2) = C/C Reference genotype = A/A

*p<0.05

Fig 1

Hazard function for recurrence of MDD by severity at first MDE.

Fig 2

Hazard function for recurrence of MDD by social avoidance.

ApaI +64978C>A was shown to significantly potentiate the effect of severity (Table 4). For A/A genotype, the hazard of recurrence increased by 6.4 times when the severity shifted from mild and moderate to severe; similarly, for the C/A genotype, the hazard ratio increased by a factor of 11.3.

Table 4

Hazard ratios for ApaI +64978C>A genotypes by severity at first MDE.

	Severity
ApaI	Mild and Moderate	Severe
A/A	0.469 (p = 0.239; 95% CI = 0.133–1.656) *	3.018 (p = 0.105, 95% CI = 0.794–14.468) *
C/A	0.289 (p = 0.033, 95% CI = 0.092–0.908) *	3.265 (p = 0.041, 95% CI = 1.047–10.182) *
C/C	1	1

*Hazard ratio, controlling for age at first MDE

Discussion

Accurate prediction and identification of factors leading to recurrence of MDD is important to improve customized management to the individual patients. However, accurate ascertainment of the diagnosis is also important before identifying those factors. A number of reviews and research articles used definitions of remission, recovery, relapse and recurrence by Frank et al. [46] but the definitions in DSM-5 [1] are clinically more pragmatic and useful, hence these were used in the current study.

From our study, the proportion of first recurrence of MDD was 20.9%. Hardeveld and colleagues reported that the rate of recurrence in specialised mental health care centres was 26.8% [47]. In another article, Hardeveld et al. reported that the rate of recurrence over a 5-year period was 60% [16]. According to the American Psychiatric Association [1], there was at least a 60% lifetime risk of recurrence after the first major depressive episode. On the other hand, the 4.3% recurrence rate reported by ten Have et al. [48] was much lower than those from this and other studies.

Our study showed that depressed individuals with a higher tendency towards social avoidance were at greater risk of recurrence of MDD. Studies regarding the influence of personality functioning on recurrence of MDD is limited. To the best of our knowledge, no previous study has reported on social avoidance as a predictor of recurrence of MDD. Empirical evidence has been accumulated to show that personality functioning is not only a modifier or a sequela of depressive disorder, but also a predictor of its incidence [49], relapses and recurrences [50, 51]. Ferster (1973) suggested that avoidance plays a role in which frequent avoidance of perceived unpleasant conditions by depressed individuals results in limited exposure to positively-reinforced behaviors and social activities [52, 53] hence increasing the risk of recurrence. In addition, anticipation of previously experienced discrimination may further cause depressed individuals to avoid participation in certain life areas, leading to greater isolation and social marginalization [54]. This finding from our study could potentially be useful for application in clinical practice; by identifying MDD patients with a high tendency to social avoidance, targeted interventions could be undertaken by the psychiatrist or clinical psychologist to mitigate the effect of this modifiable risk factor.

Similar to the results of our study, Kudo et al. (2017) did not observe a significant association between the self-criticism personality domain and recurrence of MDD [55]. However, the tools of measurement of the personality dimension (such as anxious worrying, personal reserve and perfectionism) and personality functioning (such as cooperativeness and effectiveness) in both studies were slightly different (Black Dog Institute, 2017). Individuals with a higher tendency towards neuroticism, which was not measured in our study, were being consistently reported to be associated with a higher risk of recurrence [56, 57].

Our study found that severity at first MDE increased hazard of recurrence by five and a half times. However, population and clinical studies that have been conducted used different classification systems for severity of first MDE which involved the duration of the first major depressive episode and number of symptoms present, but a more severe first MDE has consistently been shown to be related to recurrence [22]. Barkow et al. (2003) used ICD-10 criteria for severity to study the risk factors of recurrent MDE in the primary care setting and showed that severe MDD at baseline tripled the risk for development of subsequent depressive episodes [58]. NEMESIS-2, a Dutch cohort study using the Sheehan Disability Scale (SDS) to classify severity of depression, showed that severity of the last depressive episode increased recurrence of MDD by almost two times [48]. The results consistently suggested that severity of previous MDE predicted recurrence and this was also demonstrated in our study.

In terms of age at onset and gender, the results of our study replicated results of several earlier studies showing that age at onset of first MDE [17, 18, 59–60] and gender [16, 18, 59–61] did not predict subsequent episode of MDE. This is contrary to what some other studies have found [7, 11–15]. The inconsistencies in findings regarding age and gender could possibly be due to the effect of socio-cultural differences in the populations studied. In a meta-analysis of the effect of gender and age on depressive symptoms, Salk et al. put forward Eagly and Wood’s social-structural theory [62] that “larger gender differences should be observed in nations with more gender inequality”. In addition, Salk et al. found quadratic trends for gender differences in its interaction with age and age-related factors such as “stressors in adolescence and the hormonal and neurodevelopmental changes that vary by sex and peaking at ages 13–15 years and declining in the 20s”. Depression may also manifest differently in different cultures [63]. We postulate that these differences could possibly apply to recurrence, and explain the inconsistent findings between studies conducted in different countries and with different age groups studied.

Results regarding family history and recurrence also have been inconsistent–a number of previous studies have reported that a family history of psychiatric illnesses was not associated with recurrence of MDD [11, 44, 64, 65], similar to what we have found in our study. Other studies [21-23] have reported that family history significantly predicted recurrence. In a recent systematic review and meta-analysis, Buckman et al. (2018) [66] reported that “there was very limited and inconsistent evidence that family history of depression is prognostic of recurrence”. In their meta-analysis, the pooled effect size of an odds ratio of 1.36 (95% CI = 0.92, 2.01) for 3 studies was not statistically significant. The authors reported that “many studies fail to discriminate patients in their first episode from those with a history of multiple previous depressive episodes” and that “differences in the case mix across studies can lead to spurious conclusions”. This could have partially accounted for some of the inconsistencies found in the published literature.

Although ApaI +64978C>A SNP by itself is not a predictor of recurrence in the present study, we found that the interaction of ApaI +64978C>A genotype with severity at first MDE to be significant in predicting its recurrence. The hazard of recurrence for carriers of the A/A and C/A genotypes of ApaI with a severe first episode of MDD is much higher than for a less severe episode, indicating that ApaI +64978C>A polymorphisms potentiated the effect of severity on recurrence of MDD (Table 4). Kuningas et al. (32) found that Dutch patients with the baT haplotype of the Vitamin D receptor (VDR) gene were less prone to depression compared with carriers of BAt. This propensity of the A allele to increase risk of depression could possibly play a role in the interaction between ApaI polymorphism and severity. The C>A transversion at the ApaI restriction site is located in the 3’ VDR gene transcription initiation site—a ligand binding site of the VDR gene [67]—and may affect the downstream effect of the ligand-binding properties of the Vitamin D receptor, in turn affecting the Vitamin D Response Element (VDRE) complex and activation of the transcription of tryptophan hydroxylase 2 (TPH2) gene, leading to an imbalance of serotonin levels in the brain [68-70]. Although several polymorphisms, including ApaI, of the VDR gene have been described, their effects on VDR function and interaction with severity of depression are still poorly understood.

The limitations of the study include those inherent in the design of a case-control study from which data for this study was derived. Biases inherent in a case-control study include recall and selection bias. We attempted to minimize recall bias by administering the questionnaires using enumerators trained in interviewing patients. Since the questionnaires were self-administered, enumerators were on standby to ensure that any questions the patients may have were adequately addressed, with intimations from non-verbal communication from interviewers kept to a minimum. Representativeness of the sample was ensured as far as possible with the recruitment of patients who came from the catchment area of the hospital representative of the cross-section of the community as shown by their socio-demographic profiles (Table 2). The sample size undoubtedly was affected by loss to follow-up; however, there was no significant difference in the baseline profiles between those who were followed up and those lost to follow-up, indicating that the bias could have been kept to a minimum.

This study adds new information to the body of knowledge on modifiable risk factors and predictors for recurrence of MDD; future studies could further examine the role of genetic biomarkers in prognosticating recurrence in assisting health care workers in early identification and preventive management of major depressive disorder. We believe that this new finding has the potential to inform the management of MDD in which clinicians can now be alerted to potential recurrence from a fore knowledge of social avoidance and severity of a previous episode of MDD, both of which are modifiable risk factors.

Conclusion

This study suggests that social avoidance and severity of previous MDD prognosticate recurrence, with ApaI +64978C>A genotypes acting in synergy with severity in potentiating the hazard of recurrence. We suggest screening for personality factors—in particular, social avoidance; this could potentially be performed at the first major depressive episode. Psychological intervention could then be customized to focus on this modifiable factor. In addition, prompt and appropriate management of severe MDD is recommended to reduce risk of recurrence.

(DOCX)

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Genotyping of VDR Single Nucleotide Polymorphisms (SNPs).

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Genotyping for single nucleotide polymorphisms of HTR1A-rs6295 and HTR2A-rs6311.

(DOCX)

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2 Jan 2020

PONE-D-19-27517

Predictors of a first recurrence of major depressive disorder

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Comments to the Author

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Reviewer #2: Yes

**********

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Reviewer #1: No

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #2: Yes

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Reviewer #1: The authors want to identify the predictive factors for recurrence of major depressive disorders in 5-year follow-up study. They found that the social avoidance and severity of previous MDD could predict the recurrence, with 287 ApaI +64978C>A genotypes acting in synergy with severity in potentiating the risk of recurrence. It is an interesting paper, but some concerns should be addressed.

In the abstract

1. The aim of the present study should be presented.

2. “201 cases of major depressive disorder from four public hospitals in Malaysia were traced over a 5-year period from 2014 to 2017”. It should be “A total of 201 patients with major depressive disorder from four hospitals in Malaysia were followed up for 5 year”. Case should be revised as “patients” throughout the paper.

In the introduction

3. The definition of “recurrence” should cite the reference.

In the methods

4. Please clarify the inclusion and exclusion criteria for recruiting the patients.

5. Please detail the methods of DNA extraction.

6. Why the authors selected the SNPs of Vitamin D Receptor, Zinc Transporter-3, Dopamine Transporter-1, Brain-derived Neurotropic Factor, Serotonin Receptor 1A and 2A?

7. Why the author selected the variables with p values less than 0.25 from the Kaplan-Meier results, for the next Cox regression model?

In the discussion

8. The author should discuss the limitation of this study, such as the moderate sample size.

9. Overall, the language of the study should be improved, and the paper should be revised carefully.

Reviewer #2: Thank you for the opportunity to submit this manuscript. The information that Social avoidance and vitamin D receptor were prognostic of recurrence of major depressive disorder was very helpful. However, I still have several comments. I also think that some more discussion is warranted. In its current shape I am not fully convinced if this manuscript is ready to be published.

1.I don’t think that paragraph 1 with so much words is essential, considering the major aims of this manuscript.

2.I would suggest that more should be added to paragraph 3 (line 73 to 76 ) in introduction section considering the major findings of this manuscript.

3.Just as the author stated in Introduction section that those of younger age, women and family history of MDD were at higher risk of recurrence. However these findings were not consist with results of this manuscript. Readers would have great interest on the discussion of this part.

4.I would suggest that limitations should be listed in discussion section.

5.It is interesting that the interaction of ApaI +64978C>A genotype with severity at first MDE would predicted the recurrence of MDE. It would be better if more discussion about potential mechanisms added to this part(line 269 to 276) , instead of listing the result.

6.It would be better if more references from the past 5 years are added.

**********

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Reviewer #1: No

Reviewer #2: No

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15 Feb 2020

Response to Reviewers

Reviewer 1:

Abstract

• The aim of the present study should be presented.

The aim of the present study has been included. Pg 2, line 24 - 25

• “201 cases of major depressive disorder from four public hospitals in Malaysia were traced over a 5-year period from 2014 to 2017”. It should be “A total of 201 patients with major depressive disorder from four hospitals in Malaysia were followed up for 5 year”. Case should be revised as “patients” throughout the paper.

The sentence has been re-written as suggested. Pg 2, line 23 - 24

‘Case’ has been replaced with ‘patients’ where relevant.

Introduction

• The definition of “recurrence” should cite the reference.

The reference has been cited. [Reference number (1)]

It is “American Psychiatric Association. (2014). Diagnostic and statistical manual of mental disorders. 5th ed. Washington, DC: American Psychiatric Publishing”. Pg 3, line 52

Methods

• Please clarify the inclusion and exclusion criteria for recruiting the patients.

The inclusion and exclusion criteria have been added. Pg 4 – 5, lines 93 – 99

• Please detail the methods of DNA extraction.

The method of DNA extraction has been added. Pg 7, lines 148 – 156

• Why the authors selected the SNPs of Vitamin D Receptor, Zinc Transporter-3, Dopamine Transporter-1, Brain-derived Neurotropic Factor, Serotonin Receptor 1A and 2A?

Mutations in the brain-derived neurotropic factor (BDNF) [29, 30] and serotonin receptor genes HT1A and HT2A genes [31] were shown to be associated with recurrent MDD. Serotonin receptor 1A and 2A were studied in view of their close relationship with serotonin as a modifier of mood, and mood disturbances occur when there is an imbalance of serotonin metabolism caused by mutations in HT1A and HT2A genes.

In addition, findings by Lye et al. that single nucleotide polymorphisms (SNPs) of vitamin D receptor (VDR) and SLC30A3 of zinc transporter-3 (ZnT3) genes were associated with MDD [32, S1 Text], and together with evidence that SNPs of dopamine transporter-1 [33, 34] increased risk of MDD, led us to suspect if an association exists between VDR, ZnT3 SNPs and recurrence of MDD as well. Pg 4, lines 73 – 78

• Why the author selected the variables with p values less than 0.25 from the Kaplan-Meier results, for the next Cox regression model?

Statistically, Kaplan-Meier is a univariate type of analysis – it regresses the outcome (time to recurrence) on one single independent variable - hence the estimates of hazard ratios and p-values derived from each of these Kaplan-Meier runs are therefore not adjusted for confounding. So apart from selecting the obviously significant variables that had p values less than 0.05, the process of selecting those other variables with p-values greater than 0.05 but less than 0.25 is to attempt to ensure that enough leeway is given so that when we run the Cox regression model adjusting for confounders, p-values of some of those selected variables that were not significant initially with univariate analysis, (that is p-values greater than 0.05 but less than 0.25), may now fall below 0.05 after adjusting for the confounding variables in the multivariate model. A number of research articles [appended below] have used 0.25 as the cut-off point for selection of variables to enter into the Cox regression model, while Robinson (2001) provided the statistical rationale involving the cut-off point of the p-value up to a maximum of 0.25. Pg 8, lines 178 – 179

References:

1. Singh R, Tripathi V, Kalpana Singh MK, Dwivedi SN. Determinants of Birth Intervals in Tamil Nadu in India: Developing Cox Hazard Models with Validations and Predictions. Revista Colombiana de Estadística 2012; 35: 289–307.[42]

2. Sendek EM, Hebo SH. Modeling time-to-good control of hypertension using Cox proportional hazard and frailty models at Bahir-Dar Felege Hiwot Referral Hospital. Open Access Medical Statistics. 2017; 7:27–36. [43]

3. Robinson DH, Wainer H. On the Past and Future of Null Hypothesis Significance Testing. (Research Report No. RR-01-24) Statistics & Research Division Princeton, NJ, USA. Educational Testing Service. 2001; 25. [44]

Result

• The author should discuss the limitation of this study, such as the moderate sample size.

The limitations have been added. Pg 18, lines 319 - 330.

• Overall, the language of the study should be improved, and the paper should be revised carefully.

Overall, we have improved on the language, and the paper has been carefully revised according to the reviewer’s comments.

Reviewer 2:

I don’t think that paragraph 1 with so much words is essential, considering the major aims of this manuscript.

Paragraph 1 has been shortened, while paragraph 2 has been incorporated in paragraph 1. Pg 3, Lines 48 – 59

I would suggest that more should be added to paragraph 3 (line 73 to 76 ) in introduction section considering the major findings of this manuscript.

Paragraph 3, now the new paragraph 2, has been expanded, relevant to the major findings of this study. Pg 3, lines 61 – 71

Just as the author stated in Introduction section that those of younger age, women and family history of MDD were at higher risk of recurrence. However these findings were not consist with results of this manuscript. Readers would have great interest on the discussion of this part.

The findings that were inconsistent have been discussed. Pg 16 – 17, lines 277 – 301

I would suggest that limitations should be listed in discussion section.

The limitations have been included in the discussion section. Pg 18, lines 319 - 330

It is interesting that the interaction of ApaI +64978C>A genotype with severity at first MDE would predicted the recurrence of MDE. It would be better if more discussion about potential mechanisms added to this part (line 269 to 276) , instead of listing the result.

More discussion on the potential mechanisms has been added to this part. Pg 17, lines 303 – 317

It would be better if more references from the past 5 years are added.

More recent references have been added.

1. Monroe SM, Harkness KL. Recurrence in major depression: A conceptual analysis. Psychol Rev. 2011; 118: 655–674.

2. Kessler RC, Bromet EJ. The epidemiology of depression across cultures. Annu Rev Public Health. 2013; 34: 119–38.

3. Lee AS. Better outcomes for depressive disorders? Psychol Med. 2003; 33: 769–774.

4. Gueorguieva R, Chekroud AM, Krystal JH. Trajectories of relapse in randomised, placebo-controlled trials of treatment discontinuation in major depressive disorder: an individual patient-level data meta-analysis. Lancet Psy. 2017; 4:230–237.

5. Serafini G, Santi F, Gonda X, Aguglia A, Fiorillo A, Pompili M, F.Carvalhoh A, Amore M. Predictors of recurrence in a sample of 508 outpatients with major depressive disorder. J Psy Research. 2019; 114: 80-87.

6. Hardeveld F, Spijker J, De Graaf R, Nolen WA, Beekman AT. Prevalence and predictors of recurrence of major depressive disorder in the adult population. Acta Psychiatr Scand. 2010; 122(3): 184 – 191.

7. Lewinsohn PM, Zeiss AM, Duncan EM. Probability of relapse after recovery from an episode of depression. J Abnormal Psychol. 1989; 98(2): 107-116.

8. Kovac M, Obrosky DS, Sherrill J. Developmental changes in the phenomenology of depression in girls compared to boys from childhood onward. J Affect Disord. 2003;74: 33−48.

9. Lewinsohn PM, Rohde P, Seeley JR, Klein DN, Gotlib IH. Natural course of adolescent major depressive disorder in a community sample: Predictors of recurrence in young adults. Am J Psychiatry. 2000; 157:1584 – 1591.

10. Serafini G, Nebbia J, Cipriani N, Conigliaro C, Erbuto D, Pompili M, Amore M. Number of illness episodes as predictor of residual symptoms in major depressive disorder. Psychiatr Res. 2018; 262: 469–476.

11. Nierenberg AA, Husain MM, Trivedi MH, Fava M, Warden D, Wisniewski SR, Miyahara S, Rush AJ. Residual symptoms after remission of major depressive disorder with citalopram and risk of relapse: a STAR*D report. Psychol Med. 2010; 40: 41–50.

12. Nery FG, Hatch JP, Nicoletti MA, Monkul ES, Najt P, Matsuo K, Cloninger CR, Soares JC. Temperament and character traits in major depressive disorder: influence of mood state and recurrence of

episodes. Depress Anxiety. 2009;26(4):382-8

13. Nöbbelin L, Bogren M, Mattisson C, Brådvik L. Risk factors for recurrence in depression in the Lundby population, 1947–1997. J Affect Disord. 2008; 228: 125–131.

14. Wood W, Eagly AH. Biosocial construction of sex differences and similarities in behavior. Advances in Experimental Social Psychology. 2012; 46: 55–123.

15. Tsai JL, Chentsova-Dutton Y. Understanding depression across cultures. In Gotlib IH, Hammen CL, editors. Handbook of depression. New York: Guilford Press; 2002. pp. 467–491.

16. Buckman JEJ, Underwood A, Clarke K, Saunders R, Hollon SD, Fearon P, Pilling S. Risk factors for relapse and recurrence of depression in adults and how they operate: A four-phase systematic review and meta-synthesis. Clin Psychol Rev. 2018; 64:13-38.

Submitted filename: Response to Reviewers.docx

Click here for additional data file.

28 Feb 2020

Predictors of recurrence of major depressive disorder

PONE-D-19-27517R1

Dear Dr. Lye,

We are pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it complies with all outstanding technical requirements.

Within one week, you will receive an e-mail containing information on the amendments required prior to publication. When all required modifications have been addressed, you will receive a formal acceptance letter and your manuscript will proceed to our production department and be scheduled for publication.

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With kind regards,

Zezhi Li, Ph.D., M.D.

Academic Editor

PLOS ONE

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

Reviewer #2: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: (No Response)

Reviewer #2: It has clearly improved, although better it can be.I would suggest that the edior could consider to publish this manuscript .

**********

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Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No

4 Mar 2020

PONE-D-19-27517R1

Predictors of recurrence of major depressive disorder

Dear Dr. Lye:

I am pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

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