Clinicopathological Analysis And Real Time Fluorescence Quantitative PCR Detection Of Cutaneous Tuberculosis

Background:Cutaneous Tuberculosis(CTB)is a chronic infectious skin disease.The diagnosis of CTB is difficult due to the diverse clinical and pathological manifestations and low bacterial loading.Real-time fluorescence quantitative polymerase chain reaction(qPCR)has a specificity of 100% in tuberculosis(TB)and sensitivity of 24.5%-33.3% in fresh tissues of CTB.But fresh tissues are usually not retained for qPCR because of the atypical clinical manifestations of CTB,and the patients compliance of secondary sampling was poor and secondary sampling also increases the burden of patients.Therefore,MTB detection in paraffin-embedded tissues is particularly important.Previous studies confirmed that qPCR can be applied to detecte Mycobacterium tuberculosis(MTB)in paraffin-embedded tissues of systemic TB,with a sensitivity of 62.5%-74.6%.But there is till a controversy about the existence of MTB in Tuberculid.Some researchers believe that Tuberculid is an immune response of skin to blood-borne bacteria or bacterial components,but in some cases MTB was also found in skin biopsy.Objective: The aim of this study was to analyze the clinical and pathological features of CTB and Tuberculid,to improve the understanding of CTB and tuberculid;to detect MTB in fresh tissues and paraffin-embedded tissues of CTB by qPCR,and compare the sensitivity in each other;to explore the feasibility of MTB detection by qPCR in paraffin-embedded tissues of CTB;to detect MTB in Tuberculid by qPCR,to further discuss the existence of MTB in Tuberculid.Methods: Patients who were CTB and Tuberculid tentative and suspected in Department of Pathology,Shandong demartological hospital during January 2012 and March 2018 were collected.The clinical manifestations,pathological features,TB-related accessory examinations and misdiagnosis of confirmed cases were analysed.DNA was extracted from all paraffin-embedded tissues and some fresh tissues of confirmed CTB and tuberculid patients.Diagnostic kit for MTB DNA(PCR-fluorescence,Cat.#DA-052)was used,qPCR was performed using an AB 7500 fluorescence quantitative PCR system(Applied Biosystems).Results: In total,20 cases of CTB were diagnosed,including 11 cases of lupus vulgaris(LV)and 9 cases of tuberculosis verrucosa cutis(TVC);16 cases tuberculid were diagnosed,including 15 cases of Erythema induratum of Bazin(EIB)and 1 case of Papulonecrotic tuberculid(PNT).Tuberculous granuloma was found in 36.4% of LV,22.2% of TVC,and 6.7% of EIB.The qPCR sensitivity for fresh CTB tissue samples and paraffin-embedded tissue samples were 50% and 35%,respectively.MTB was not detected in both fresh and paraffin-embedded tissues of tuberculid.Conclusion: The clinical manifestations of CTB are various.The rate of misdiagnosis and delayed diagnosis is high.30% cases can be diagnosed by histopathology,and 70% cases can be suggested by histopathology.QPCR can be used to detect MTB in fresh tissues and paraffin-embedded tissues of CTB.QPCR can assist the early diagnosis of CTB.Compared with traditional techniques acid-fast staining,qPCR detection has a higher sensitivity.In clinical work,if CTB is considered in the first diagnosis,we suggest taking fresh tissues for qPCR in the same time as pathological samples taken.If the clinical manifestations are atypical,testing paraffin-embedded tissues by qPCR is also an important way to assist etiological diagnosis when pathological results indicate the possibility of CTB.Tuberculid may be the skin's immune response to blood-borne bacteria or bacterial components,no MTB was found in Tuberculid lesions.