Table 4.

Advantages and disadvantages of different molecular methods

S. no.	Assay	Types of leishmaniasis	Specificity	Sensitivity	Advantage	Disadvantage	References
1.	Conventional PCR	Cutaneous leishmaniasis (CL)	87.61–100%	100%	Precise results, high specificity, and sensitivity. Uncomplicated Diagnostic interpretations	Time consuming and incompetent to evaluate the destined DNA. Qualitative Approach. Restricted detection range	Moreira et al. (2007), Zeyrek et al. (2018), Abd El-Salam et al. (2014), De Paiva-Cavalcanti et al. (2015)
2.	Nested PCR	Cutaneous Leishmaniasis (CL), Visceral Leishmaniasis (VL)	90–100%	–	Shows higher sensitivity and specificity. A convenient method for investigating the molecular epidemiology in the field	Qualitative test. Incompetent to evaluate the target DNA requires prolonged time and is expensive	Shirian et al. (2014), Oliva et al. (2006), De Paiva-Cavalcanti et al. (2015)
3.	Real Time-PCR	PKDL, VL	91.2–93.33%	100%	Elevated specificity and sensitivity, Numerical potential and rapid results. Differentiation of species can be achieved by melting Temperature	Complexity in elucidating the outcomes, Requires a skilled operator Presence of thermocycler makes it costly	Hossain et al. (2017, Ghosh et al. (2018), De Paiva-Cavalcanti et al. (2015)
4.	NASBA	VL	93.3–97.5%	100%	NASBA is the only isothermal amplification method that utilizes RNA as starting material. There is no requirement of the complicated laboratory structure. Shows higher specificity and rapid results	Prone to contamination of ribonuclease, which can degrade the target RNA	van der Meide et al. (2005), Mugasa et al. (2010), Zanoli and Spoto (2013), De Paiva-Cavalcanti et al. (2015)