One hundred and forty five (145) South Indian Tamil patients who had a diagnosis of Bipolar Disorder according to DSM-IV criteria (American Psychiatric Association) were enrolled for the present study [7]. The patients were recruited from both out-patient and in-patient services of the Department of Psychiatry at Jawaharlal Institute of Postgraduate Medical Education and Research (JIPMER), Puducherry, India, a major tertiary referral centre in South India between September 2012 and April 2016. Clinical history was obtained for each patient through structured interviews conducted by trained psychiatrists. All subjects were screened before inclusion for any recent infections and existing inflammatory conditions or ongoing treatment with immunomodulatory drugs. Psychiatric evaluations were performed to assess the severity of symptoms and level of functioning of the subjects. Severity of manic symptoms was rated with the Young’s Mania Rating Scale (YMRS) [8] and depressive symptoms with the Montgomery-Asberg Depression Rating Scale (MADRS) [9]. Remission was defined as a score of <7 on YMRS, and <10 on MADRS. One hundred and fifty one (151) Healthy Control (HC) subjects (age, sex, and ethnicity matched) without a personal or family history (first degree) of psychiatric, neurological or autoimmune disorders were also enrolled from volunteers among hospital staff and the general public. All participants provided written informed consent for participation in the study which had previously been reviewed and approved by the institutional ethics committee.

CRP status was profiled in these subjects by qualitative latex agglutination method using “CRP Latex” kits (Reckon Diagnostics, Vadodara, India) according to the manufacturer’s protocol. The commercial kit had a sensitivity of 0.6 mg/dL.

The sample size for the study which had 80% power and 95% confidence interval was determined using the Sample Size for Unmatched Case-Control Studies (SSCC) module of the OpenEpi V3 open source calculator. The frequency of CRP positivity, defined as >0.6 mg/dL serum, was compared between BD and HC control groups using the chi-square test. The Odds Ratio (OR) with 95% confidence interval was determined. A (two tailed) p-value <0.05 after correction was regarded as statistically significant. Sensitivity, specificity, positive and negative predictive values were determined. GraphPad prism V7 was used to perform the statistical tests. Logistic regression analysis to control for potential confounders including age, gender, BMI, and smoking status was carried out using Statistical Package of Social Sciences (SPSS) Version 20.0.

The demographic details of the subjects are presented in [Table/Fig-1]. Overall, the mean age of patients and controls were 32.95 (±10.57) and 33.12 (±11.45) years respectively. Gender distribution was not significantly different between the patient and control groups [Table/Fig-1]. The mean BMI of the patient group was 24.17 (±4.34) and 16% of the patients were active smokers.

CRP positivity was significantly higher in BD patients (p=0.0001, OR=4.3, 95% CI=2.0-9.4) compared to controls [Table/Fig-2].

For the purpose of analysis, the patients who had residual symptoms were grouped with the patients in remission at the time of enrolment. Upon comparing subgroups of cases who were in acute mood episode at enrolment with those who were in remission/residual phase, it was found that CRP positive status was significantly higher in the acute disease group (p=0.003, RR=1.48, 95% CI=1.2-1.8) when compared to subjects in remission or residual disease [Table/Fig-3], and remained significant after controlling for confounders namely age, gender, BMI and smoking status of the subjects.

Further, upon comparing among patients in acute phase, It was found that CRP status did not differ significantly between patients in an acute manic episode (n=85) and in acute depression (n=9). However, CRP positive status was significantly higher in both acute mania (p=0.01, OR=4.10, 95% CI=1.33-12.71) and acute depression (p=0.01, OR=9.4, 95% CI=1.8-49.7) when compared to subjects in remission or residual disease [Table/Fig-4].

The sensitivity of the test for distinguishing BD cases and healthy controls was 21.4% (95% CI=0.15-0.29) and specificity was 94.04% (95% CI=0.89-0.97); the positive and negative predictive values were 77.5% (95% CI=0.63-0.88) and 55.5% (95% CI=0.49-0.61) respectively. As for distinguishing acute mood episodes from remission/residual phase the sensitivity and specificity were 29.7% (95% CI=0.21-0.39) and 92.2 (95% CI=0.81-0.96) and the positive and negative predictive values were 87.1% (95% CI=0.71-0.95) and 41.2% (95% CI=0.33-0.50) respectively.

Immune related mechanisms are associated with the core psychopathology of BD [3] and a chronic low-grade inflammation is well admitted to be operative at least in a subset of BD patients, especially in those with more severe symptoms [2]. CRP, an acute phase reactant produced by the liver and secreted into the blood, is a marker of inflammation used quite often in the clinical settings. It has been shown that CRP association is not affected by psychotropic medication [2], increasing the utility of CRP as a marker if validated.

While in some studies high CRP levels has been shown to be significantly associated with acute mood states, especially mania [1,2,4], other studies did not demonstrate significant differences in CRP levels based on the mood states [3,5] although levels were higher compared to healthy controls. The differences in the findings could be attributed to variation in study design, relatively small sample sizes and not controlling for cofounders that could potentially influence CRP levels like BMI, smoking status etc. Also, medication, lifestyle and genetic factors could likely contribute to the confounding effect on the observations. In addition, in one study, the authors suggest that some of the patients could have had a shift in affective status between the time of clinical assessment and blood sampling as both were not carried out on the same day as done in the present study. Such confounders, it is suggested, could introduce random noise thereby reducing the likelihood of finding associations [3]. CRP alterations across mood states have also been demonstrated by prospective longitudinal studies [10]. The present study also did not distinguish acute phase mania from acute depression with respect to CRP status, although CRP status among patients with both acute mania and acute depression were significantly different from patients in remission/ having residual symptoms. Yet, the lack of association in the present study should be addressed with caution, owing to the small sample sizes compared after subgrouping.

An earlier meta-analysis showed that apart from the acute mood states, elevated CRP was also observed in euthymic BD patients compared to healthy controls [6]. However, in the present study, no significant differences in CRP status were observed between euthymic BD and healthy controls.

As bipolar disorder is clearly associated with higher risk of metabolic syndrome [11] and conceived as a multi-system inflammatory disease [12], CRP has been demonstrated to be useful as a marker of metabolic syndrome in bipolar disorder patients [13] and thus of higher load of cardiovascular comorbidities.

A previous longitudinal study with a small sample of BD patients on treatment could not distinguish CRP level-based BD subgroups, and reported no statistically significant association between higher CRP values and a more unfavourable outcome in either euthymic or non-euthymic patients [14]. In the present study, upon cross-sectional examination, it was found that, significant difference in the CRP status between patients in an acute episode and those in remission/ residual status, after controlling for potential confounders.

The major aim of present study was to develop and validate CRP status as marker for impending acute episodes in BD patients, especially dedicated to our underserved poor socio-economic patient populations. The method of CRP testing used herein was simple and does not require much expertise, equipment, or sophisticated laboratory settings to carry out. It is also cost effective as the costs per test are as low as 0.2 USD (approx.).

The approach herein validated in a cross sectional design, although warranting further longitudinal validation, was simple, relatively sensitive, specific and cost-effective and easy to implement in point-of care-out-patient, bed side, or field psychiatric consultation settings without the requirement of a laboratory.

Financial support: This work was supported by a research grant from the Centre Franco-Indien pour la Promotion de la Recherche Avancée (CEFIPRA), Project No. 5103-I. Aparna Sundaresh gratefully acknowledges the receipt of Intramural Funding for PhD work from Jawaharlal Institute of Postgraduate Medical Education and Research (JIPMER).

Other interest: The work presented here was part of a PhD thesis. The ‘primary researcher’ is Aparna Sundaresh, and the ‘Supervisor’ is Dr. Vir Singh Negi.