Diagnostic performance of measuring antibodies to the glycopeptidolipid core antigen specific to Mycobacterium avium complex in patients with rheumatoid arthritis: results from a cross-sectional observational study

A cross-sectional observational study was conducted. The study sample consisted of 396 patients who were treated for RA between May and October 2013 at the Hirose Clinic of Rheumatology, which is an outpatient clinic located in Saitama prefecture in Japan. The inclusion criteria for patients were fulfillment of the 2010 American College of Rheumatology/European League against Rheumatism classification criteria for RA [11]. Patients younger than 20 years of age and those who had already been enrolled in a clinical study with the intervention of a study drug were excluded. A total of 824 patients with RA visited our clinic during the study period. We randomly screened a total of 443 of 824 patients with RA, and a total of 381 patients with RA (46.2 %) were not screened.

Sera from all participants were screened for anti-GPL IgA antibodies, and all the participants underwent chest radiography. For statistical analyses, recorded data, including demographics, disease activity, comorbidities, treatments, and laboratory data, were obtained at the time of consent. All participants provided written informed consent before study enrollment according to the Declaration of Helsinki (World Medical Association General Assembly, October 2008). This study was conducted with the approval of the Hattori Clinic Institutional Review Board (Hachioji, Tokyo, Japan), which was responsible for reviewing and approving the study.

We defined MAC-PD according to the 2007 diagnostic criteria for NTM lung disease proposed by the American Thoracic Society and Infectious Diseases Society of America [1]. Both of the following criteria had to be met clinically: (1) pulmonary symptoms, nodular or cavitary opacities on chest radiographs, or HRCT scans manifesting multifocal bronchiectasis with multiple small nodules; and (2) appropriate exclusion of other diagnoses. Only one of the following criteria was required microbiologically: (1) positive culture results for at least two separate expectorated sputum samples, (2) positive culture results for at least one bronchial wash or lavage, or (3) transbronchial or other lung biopsy specimens with mycobacterial histopathological features. A diagnosis of MAC-PD infection required the fulfillment of the clinical and microbiological criteria described above.

All serum samples were sent to the Life Science Institute, Inc. (Tokyo, Japan), where a commercially available EIA kit used to detect serum IgA antibodies specific to the MAC-GPL core antigen (Capilia MAC Antibody ELISA; TAUNS Laboratories, Shizuoka, Japan) was employed with a previously described method [6]. The GPL core antigen is a major cell surface antigen in MAC that is absent on the cell wall of either M. tuberculosis or Mycobacterium kansasii [10]. At the Life Science Institute, the cutoff value was defined as 0.7 U/ml according to previous findings [6].

Chest x-rays were assessed by consensus reading by four rheumatologists and one respiratory physician (WH, KI, MM, TN, TU). All patients with RA who had abnormal shadows on their chest x-rays underwent chest CT. Chest CT images from patients with RA were reviewed with consensus reading by one rheumatologist and one respiratory physician experienced in CT (WH, TU). To localize the infection by MAC, the lungs of each patient were divided into 10 fields (right lung: S1 + S2, S3, S4 + S5, S6, and S7 + S8 + S9 + S10; left lung: S1 + S2, S3, S4 + S5, S6, and S8 + S9 + S10) according to Moore’s definition [12]. Each field was scored with reference to the presence of bronchiectasis, centrilobular nodules, air space disease, a cavity, and nodules larger than 5 mm as described in a previous study [13]. The severity of bronchiectasis was categorized as grade 1 (diameter of the bronchus less than twice as large as the accompanying vessel), grade 2 (diameter of the bronchus at least twice as large as the accompanying vessel), or grade 3 (cystic bronchiectasis). The distribution of centrilobular nodules (size less than 5 mm) was categorized as grade 1 (0–50 % of the segment), grade 2 (50–100 % of the segment), or grade 3 (100 % of the segment). Air space disease was defined as an area of patchy or dense consolidation. Air space disease and cavity formation were categorized as grade 3 on the basis of a previous study in which these two findings were closely related to clinical features [14]. Nodular sizes larger than 5 mm were also scored as grade 1. Grades of the 10 fields were summed to calculate scores for these radiological findings. Thus, the maximum scores of bronchiectasis, centrilobular nodules, air space disease, cavities, and nodules larger than 5 mm in both lungs were 30, 30, 30, 30, and 10, respectively. Scores for the extent of MAC-PD were added to give total CT scores [13]. Thus, the maximum possible total CT score in both lungs was 130.

All patients with RA with abnormal findings on chest CT compatible with MAC-PD submitted expectorated sputum for examination on three consecutive days. An early-morning specimen was collected in a sterile cup, and it was immediately transported to the microbiology laboratory at the Life Science Institute. The sputum was examined and cultured for mycobacteria. Identification of MAC was confirmed by TaqMan polymerase chain reaction (Applied Biosystems, Foster City, CA, USA) at the same laboratory. The patients whose expectorated sputum was negative for MAC were recommended to undergo bronchoscopy at the Fukujuji Hospital, which was performed once the patient’s consent was obtained.

All data were analyzed using SAS software version 9.3 (SAS Institute, Cary, NC, USA). Demographics and clinical characteristics were expressed as means and standard deviations for continuous variables or as frequencies and percentages for categorical variables. Comparisons between patients with RA with and without MAC-PD were made using Student’s t test for continuous variables. Pearson’s χ² test was used for categorized variables. All P values were two-tailed, and P values less than 0.05 were considered significant. The Mann–Whitney U test was used to compare the differences in titers of anti-GPL antibodies between patients with and those without MAC organisms in bronchoalveolar lavage fluid (BALF) samples. Correlation coefficients between the extent of MAC-PD on chest CT images and titers of anti-GPL IgA antibodies were analyzed using a simple regression analysis.