From time immemorial, leprosy is a grossly mutilating disease associated with social stigma and taboos, particularly in underdeveloped nations. The global caseload of leprosy has reduced by almost 90% over the last 20 years and 15 million cases have been detected and cured worldwide. Three hundred thousand (0.3 million) new cases were detected during the year 2005 [1]. The principal factor contributing to this worldwide success is attributed to the introduction of standardized MDT regimens against the causative agent, Mycobacterium leprae. Further, leprosy elimination campaigns for case detection in communities, training of physicians and leprosy health care workers, community awareness towards prevention and control of leprosy have also proven to be beneficial. Elimination is defined as a reduction in the prevalence of leprosy patients receiving antibacterial therapy to less than 1 per 10,000 populations[2], which indicates that the disease is no longer considered a major public health problem. India has achieved the prevalence rate of less than 1 per 10,000 populations in 2006. But the incidence rate remains high in six countries of South East Asian region including India. India alone accounted for 60% of the world's newly detected cases [1, 3]. This might be due to lack of consistent information on the core elements of this infectious disease, e.g. source of infection, reservoir and mode of transmission, host factors related to immunity of disease etc [4–6]. It has been observed that though prevalence has declined since initiation of MDT, the incidence has not shown a similar decline during the same period i.e. after implementation of MDT [7–10].

Leprosy, particularly MB type, is highly contagious and infectious may spread to several contacts of patients even before clinical diagnosis. In case of MB leprosy, contacts staying in the same house are at higher risk of getting infection compared to contacts staying at the neighboring houses [10–14]. Therefore, early detection of infections among close contacts followed by effective chemotherapy is likely to bring down the spread of disease leading to a decline in the overall incidence rate. Unfortunately, the conventional method of contact tracing fails to detect fresh cases before it becomes transmissible to others (person to person transmission) [15] Hence, it is necessary to have an alternative and more effective tool for an early detection to prevent and control further transmission.

Previous work carried out by us, has led to the development of a Multiplex PCR (M-PCR) for early diagnosis of leprosy. The technique was standardized and was evaluated with high sensitivity and specificity [16].

The present study was carried out to evaluate whether the same technique could be used as a better diagnostic tool for early detection of leprosy cases and contacts for prediction of future cases of leprosy.

From the above table it is clear, that on the whole M-PCR can detect a higher number (79.2% to 89.4%) of leprosy cases compared to SSS test (19.1% to 87.5%). The sensitivity of M-PCR in case of tuberculoid type is higher than smear examination. It is noted that the positivity was considerably higher in tuberculoid cases (TT 79%, BT 81.9%) in comparison to SSS (TT 31.4%, BT 19.1%). Multiplex PCR of nasal swab was found to be positive 49.4% (42/85), 51.6% (32/62), 62.5% (25/40) and 78.1% (75/96) in the categories of TT, BT, BL and LL respectively. This indicates that nasal swab could be an alternative, non-invasive procedure for M- PCR for leprosy detection. PCR results were confirmed by sequencing. (Figure -1)

The data obtained from M-PCR of nasal swab of 110 household contacts of MB patients and 72 of PB patients are presented in table 2. The result shows that 11.7% adults and 9.5% children among MB contacts and 2% adult among PB contacts showed evidence of M leprae DNA in their nasal swabs. These contacts were followed up prospectively for two years since the day of collection of nasal swabs. It was observed that 1 adult and 1 child developed leprosy from M-PCR positive MB contacts during the two-year observation period. None of the contacts from PB cases developed clinical leprosy during the follow up period (table-2)

Multibacillary leprosy contact			Paucibacillary leprosy contact
Number of persons tested with M-PCR (Nasal Swab)	Number positive (%)	Development of leprosy cases among contacts within 2 year follow up	Number of persons tested with M-PCR (Nasal Swab)	Number positive (%)	Development of leprosy cases among contacts within 2 year follow up
Adult = 68	8 (11.7%)	1	Adult = 48	1 (2%)	0
Child = 42	4 (9.5%)	1	Child = 24	0 (0%)	0
Total = 110	12 (10.9%)	2	Total = 72	1 (1.3%)	0

Leprosy is a disease with long incubation period and the symptoms are difficult to perceive at the early stage of infection. Self-healing does occur in a large number of infected cases. Moreover many patients with early signs are not aware that they are suffering from leprosy and thus clinical diagnosis is often delayed. Clinical diagnosis is possible only when the patient is symptomatic, exhibiting lesions and satisfies the criteria of cardinal signs of leprosy [23]. A sizable proportion of new cases are among children (WHO 2004),[21] as they often remain in close contact with infected family members sharing same dwelling units that facilitate infection in them [22] Similarly, an infected child could pass this infection to other children while they are in contact for longer duration, i.e, playing in a group or in school. This often goes un-noticed during initial phase because of its slow and silent nature of early transmission.

Another important and effective strategy for reduction of incidence of leprosy is contact tracing and early detection of cases among them. In our pilot study M-PCR technique was applied in the leprosy contacts for adopting a better way of community-based early case detection. Contacts of PB and MB cases both were followed up after testing their nasal swabs by M-PCR. It was noted that two of the twelve M-PCR positive cases developed as clinical cases and none of the M- PCR negative cases developed any form of leprosy during this period. It is interesting to note that a significantly larger numbers (12 numbers) of MB contacts were positive by M-PCR than that of PB contacts (2 numbers) who did not develop leprosy during the follow up period.

It has been estimated that 6-8% of household contacts develop clinical symptoms of leprosy within two years of follow up since the diagnosis of the index cases[24]. In our study 10.9% MB contacts were PCR positive and 1.8% developed the disease within two years of study period. This low number of detection in our study group could be due to the lower transmission dynamics in the community during the post elimination era. Keeping in mind the long incubation period of the disease it is imperative that the contacts need to be followed and observed for a longer period.

The trend that we find in our study can serve as important clue that the contacts of MB patients are at increased risk of developing leprosy in the future. This hypothesis needs to be tested on large scale of population over a long interval before coming to any conclusion, since the authors have no intention to put forth the MB patients into social ostracism under the fear of infection. Whatever the relationship between positivity of PCR and development of the disease, PCR is much more sensitive than microscopic examination for direct detection of bacilli [25, 26]. Moreover, at present there is no other more sensitive alternative tool available for early detection of leprosy other than PCR and serological test [22]. Present M-PCR test has been standardized based on the presence of M. leprae DNA which does not support the presence of viable organisms. Effort should be made to standardize a method based on mRNA detection so that viability of M. leprae bacilli could also be ascertained [27–30].

It should be also noted that the PCR positivity in contact persons indicate the presence of M. leprae DNA only and not infection and therefore, a careful follow up of them should be done and treatment should be started immediately after the development of first sign and symptoms of leprosy.

This study indicates that M-PCR can be used as an efficient tool for early detection of leprosy among contacts. However it needs further in-depth study with adequate population size and controls over a long period of time. As this M- PCR can only detect the presence of M. leprae DNA, hence a contact with positive PCR result must be followed up regularly for detection of any development of disease. As leprosy has a long and varied incubation period, a long term follow up/observation is necessary to establish clearly the early case detection efficiency of M- PCR.

This M- PCR is a relatively expensive procedure compared to other methods of detection of infection caused by M. leprae. It has its limitations in the viability detection of the bacteria. In addition, it also requires a well-equipped laboratory which is difficult to replicate in the field. When tests are performed in bulk, the cost is expected to go down significantly. Hence, we recommend its application in a large number of samples to make it cost effective. The test needs to be evaluated further as it could serve as a better diagnostic tool for early case detection and their treatment to achieve faster control of leprosy.