Improved Glass Bead-Vortex Oscillation Method for DNA Extraction from Diatom / 法医学杂志

OBJECTIVES@#To examine the effect of improving diatom DNA extraction by glass bead - vortex oscillation method.@*METHODS@#The DNeasy PowerSoil Pro kit was used as control, two plant DNA extraction kits with different principles (New Plant genomic DNA extraction kit and Plant DNA Isolation kit) and one whole blood DNA extraction kit (whole blood genomic DNA extraction kit) were selected to extract diatom DNA from lung tissue and water sample of the same drowning case. The combination of mass ratio of glass beads with different sizes and vortex oscillation time was designed, and the optimal DNA extraction conditions were selected with the addition of glass beads oscillation. The extracted products of the conventional group and the modified group were directly electrophoretic and detected by diatom specific PCR. Finally, all the extracts were quantified by qPCR, and the Ct values of different groups were statistically analyzed.@*RESULTS@#When the frequency of vortex oscillation was 3 000 r/min, the optimal combination of DNA extraction was vortex oscillation for 4 min, and the mass ratio of large glass beads to small glass beads was 1∶1. The DNeasy PowerSoil Pro kit was used as a reference, and the Ct value of 10 mL water sample was greater than that of 0.5 g tissue. The Ct values of the other three kits used for plant DNA extraction decreased after the glass beads-vortex oscillation method was used, and the Ct values of the tissues before and after the improvement were statistically significant (P<0.05). The whole blood genomic DNA extraction kit used in this study could successfully extract diatom DNA, the extraction of water samples was close to DNeasy PowerSoil Pro kit, after the modified method was applied to tissue samples, the difference in Ct value was statistically significant (P<0.05). However, when the three kits were used to extract diatom DNA from water samples, Ct values before and after the improvement were only statistically significant in New Plant genomic DNA extraction kit group (P<0.05).@*CONCLUSIONS@#The improved glass bead-vortex oscillation method can improve the extraction efficiency of diatom DNA from forensic materials, especially from tissue samples, by plant and blood DNA extraction kits.

Application of Quantitative Analysis of Diatoms in Lung Tissue for the Diagnosis of Drowning of Experimental Animals / 法医学杂志

Objective To discuss the application value of diatom examination in lung tissue for the forensic diagnosis of drowning. Methods The experimental animals were divided randomly into drowning, postmortem submergence and dying on land group. Diatoms in lung tissue and drowning fluid were analyzed quantitatively by microwave digestion-vacuum filtration-automated scanning electron microscopy diatom examination method. The ratios of content of diatoms in lung tissue and drowning fluid （CL/CD ratio） were recorded. Results The CL/CD ratios of experimental rabbits in the drowning group （5.82±3.50） were much higher than that of postmortem submergence group （0.47±0.35）; the CL/CD ratios of different parts of the lung lobes of experimental pigs in the drowning group were higher than that of postmortem submergence group （P<0.05）; in seawater, brackish water, river fresh water and lake fresh water, the CL/CD ratios of experimental pigs in the drowning group were higher than that of postmortem submergence group （P<0.05）. In animal experiments, all the cases with CL/CD ratio >1.6 were from drowning group. Conclusion CL/CD ratio is an indicator with good application prospects in the diagnosis of drowning.

Research Progress on Determination of Drowning Site / 法医学杂志

The bodies found in water are one of the most common types in forensic practice. The dis-covery site of the body is often not the drowning site. However, the determination of drowning site is vital for the identification of victim. Inorganic particles and planktons, such as granular impurities, diatoms and bacteria, are valuable markers for the diagnosis of drowning. By comparing the granular impurities and planktons in tissues and suspicious drowning mediums, the drowning site can be concluded based on their similarity of types and distribution, which has practical applied value. In this paper, the research progress on determination of drowning site is summarized to provide reference for the peers.

Value of specific 16S rDNA fragment of algae in diagnosis of drowning: an experiment with rabbits / 南方医科大学学报

<p><b>OBJECTIVE</b>To establish a method for amplifying specific 16S rDNA fragment of algae related with drowning and test its value in drowning diagnosis.</p><p><b>METHODS</b>Thirty-five rabbits were randomly divided into 3 the drowning group (n=15), postmortem water immersion group (n=15, subjected to air embolism before seawater immersion), and control group(n=5, with air embolism only). Twenty samples of the liver tissues from human corpses found in water were also used, including 14 diatom-positive and 6 diatom-negative samples identified by microwave digestion-vacuum filtration-automated scanning electron microscopy (MD-VF-Auto SEM). Seven known species of algae served as the control algae (Melosira sp, Nitzschia sp, Synedra sp, Navicula sp, Microcystis sp, Cyclotella meneghiniana, and Chlorella sp). The total DNA was extracted from the tissues and algae to amplify the specific fragment of algae followed by 8% polyacrylamide gelelectrophoresis and sliver-staining.</p><p><b>RESULTS</b>In the drowning group, algae was detected in the lungs (100%), liver (86%), and kidney (86%); algae was detected in the lungs in 2 rabbits in the postmortem group (13%) and none in the control group. The positivity rates of algae were significantly higher in the drowning group than in the postmortem group (P<0.05). Of the 20 tissue samples from human corps found in water, 15 were found positive for algae, including sample that had been identified as diatom-negative by MD-VF-Auto SEM. All the 7 control algae samples yielded positive results in PCR.</p><p><b>CONCLUSIONS</b>The PCR-based method has a high sensitivity in algae detection for drowning diagnosis and allows simultaneous detection of multiple algae species related with drowning.</p>

Diatoms in drowned and postmortem immersed rabbits' lungs / 法医学杂志

OBJECTIVE@#To investigate the numbers, sizes and types distribution of diatoms in drowned and postmortem immersed rabbits' lungs.@*METHODS@#Sixty-two rabbits were randomly divided into drowning group (n = 30), postmortem immersion group (n = 30) and land death group (n=2), and the diatoms in each lung lobe were analyzed quantitatively and qualitatively by microwave digestion and scanning electron microscopy.@*RESULTS@#In the drowning group, the diatoms were detected in each lung lobe with Cyclotella and Melosira in the majority. In the postmortem immersion group, Cyclotella was in the majority. And the diatoms weren't detected in some lung lobes in postmortem immersion. There were significant differences in the detection rates of upper lobe of left lung, middle lobe and cardiac lobe of right lung in two groups (P < 0.05).@*CONCLUSION@#Based on the microwave digestion and scanning electron microscopy, the numbers, sizes and types distribution of diatoms in drowned and postmortem immersed rabbits' lungs can be analyzed and used as references for testing theory.

Progress in application of microbeam X-ray fluorescence spectroscopy in forensic science / 法医学杂志

Microbeam X-ray fluorescence (micro-XRF) spectrometry has been raised as an analytical technique of microbeam during the recent years. With its advantages of high sensitivity, small sample requirement, high testing accuracy and non-destruction, the technique is widely utilized in forensic science. This review bases on recent researches at home and abroad, describes its applications including identification of gunshot residue, visualization of fingerprints, discrimination of drug source, production process, and other material evidences of analysis in crime scene. Thanks to the advances in technology, intelligent and portable micro-XRF equipment has appeared to be applied. It is believed that it may be more popular and frequent in administration of forensic science in the near future.

Determination of acid-resistant silicic particles in lung by micro-XRF and its application in diagnosis of drowning / 法医学杂志

OBJECTIVE@#Using microbeam X-ray fluorescence (Micro-XRF) analyzer for determination of acid-resistant silicic particles in lung, and to explore its potential application in diagnosis of drowning.@*METHODS@#Thirty two white rabbits were divided randomly into drowning group (n=12), post-mortem immersion group (n=10) and control group (n=10). Lungs and water sample were collected for determination of area concentration of acid-resistant silicic particles using Micro-XRF method.@*RESULTS@#The area concentration of acid-resistant silicic particles for the drowning water sample was 4.4 mm2/mL. For the lungs of drowning group, the post-mortem immersion group and the control group, the determined average values were (25.30 +/- 10.95) mm2/g, (1.68 +/- 0.63) mm2/g and (1.65 +/- 0.85) mm2/g, respectively, with a statistically significant difference between the drowning group and the other two groups.@*CONCLUSION@#The area concentration of acid-resistant silicic particles in lungs may be used as an indicator of drowning. The method is highly sensitive and rapid. It provides a potential application in drowning diagnosis.