Aim: Isolation, dark field detection and microscopic agglutination test (MAT) are considered ―gold standard‖ tests for diagnosis of Leptospirosis. Several PCR assays are reported but very few have been evaluated for detection of Leptospirosis. Therefore, this study was undertaken. This study aims to design and standardize polymerase chain reaction (PCR) -based DNA sequencing technique for the detection of pathogenic Leptospira from peripheral blood of patients clinically diagnosed with septicemia. Methodology and Results: Two hundred and seven (207) blood samples from patients were diagnosed with septicemia which includes 100 bacterial (other than Leptospira) culture positive and 107 bacterial culture negative samples were studied. Primers for Nested PCR targeting LipL32 gene of Leptospira interrogans were designed and the specificity of primers was tested against serum samples positive/negative by either MAT or dark field microscopy. PCR amplified products were further confirmed by DNA sequencing. The standardized nPCR was sensitive and specific to Leptospira interrogans. Twenty-one (21%) out of 100 culture positive blood samples, three (2.8%) out of 107 culture negative samples showed nPCR positivity and were confirmed as Leptospira interrogans by DNA sequencing (p<0.001). A sensitive nPCR specific to Leptospira interrogans was developed. Conclusion, significance and impact of study: The p value (<0.001) signifies that Leptospira is commonly associated with other bacteria circulating in blood indicating that a decreased immune status is created primarily by a bacterium with enhanced possibility of development of Leptospiral infection probably be of an endogenous origin.

Background: Evaluate morbidities and “quality” of fiducial marker placement in primary liver tumours (hepatocellular carcinoma [HCC]) for CyberKnife. Materials and Methods: Thirty-six HCC with portal vein thrombosis(PVT) were evaluated for “quality” of fiducial placement, placement time, pain score, complications, recovery time and factors influencing placement. Results: One hundred eight fiducials were placed in 36 patients. Fiducial placement radiation oncologist score was “good” in 24(67%), “fair” in 4(11%), and “poor” in 3(8%) patients. Concordance with radiologist score in “poor”, “fair”, and “good” score was 2/2(100%), 4/5(80%), and 24/27(89%), respectively(p=0.001). Child-Pugh score(p=0.080), performance status(PS) (p=0.014) and accrued during “learning curve”(p=0.013) affected placement score. Mean placement time(p=0.055), recovery time(p=0.025) was longer and higher major complications(p=0.009) with poor PS. Liver segment involved(p=0.484) and the Barcelona Clinic Liver Cancer(BCLC) stage did not influence placement score. “Good” placement score was 30% in first cohort whereas 93% in last cohort(p=0.023). Time for placement was 42.2 and 14.3 minutes, respectively(p=0.069). Post-fiducial pain score 0–1 in 26 patients(72%) and pain score 3–4 was in 2(6%). Five patients (14%) admitted in “day-care”(2 mild pneumothorax, 3 pain). Mortality in 1 patient(3%) admitted for hemothorax. Conclusion Fiducial placement is safe and in experienced hands, “quality” of placement is “good” in majority. Major complications and admission after fiducial placement are rare. Complications, fiducial placement time, recovery time is more during the “learning curve”. Poor Child-Pugh score, extensive liver involvement, poor PS have higher probability of complications.

Background: Evaluate morbidities and “quality” of fiducial marker placement in primary liver tumours (hepatocellular carcinoma [HCC]) for CyberKnife. Materials and Methods: Thirty-six HCC with portal vein thrombosis(PVT) were evaluated for “quality” of fiducial placement, placement time, pain score, complications, recovery time and factors influencing placement. Results: One hundred eight fiducials were placed in 36 patients. Fiducial placement radiation oncologist score was “good” in 24(67%), “fair” in 4(11%), and “poor” in 3(8%) patients. Concordance with radiologist score in “poor”, “fair”, and “good” score was 2/2(100%), 4/5(80%), and 24/27(89%), respectively(p=0.001). Child-Pugh score(p=0.080), performance status(PS) (p=0.014) and accrued during “learning curve”(p=0.013) affected placement score. Mean placement time(p=0.055), recovery time(p=0.025) was longer and higher major complications(p=0.009) with poor PS. Liver segment involved(p=0.484) and the Barcelona Clinic Liver Cancer(BCLC) stage did not influence placement score. “Good” placement score was 30% in first cohort whereas 93% in last cohort(p=0.023). Time for placement was 42.2 and 14.3 minutes, respectively(p=0.069). Post-fiducial pain score 0–1 in 26 patients(72%) and pain score 3–4 was in 2(6%). Five patients (14%) admitted in “day-care”(2 mild pneumothorax, 3 pain). Mortality in 1 patient(3%) admitted for hemothorax. Conclusion Fiducial placement is safe and in experienced hands, “quality” of placement is “good” in majority. Major complications and admission after fiducial placement are rare. Complications, fiducial placement time, recovery time is more during the “learning curve”. Poor Child-Pugh score, extensive liver involvement, poor PS have higher probability of complications.