A method for bone marrow extraction of diatoms for forensic science applications

Diatoms show potential as trace evidence indicators, particularly as evidence of drowning. Often, the diatom test to diagnose drowning is done on soft tissue or bone marrow from a recently deceased individual. This method presented here combines elements from previous forensic literature and methods of diatom isolation in phycology to extract diatoms from bone marrow of skeletal remains for forensic use. This diatom extraction method is time-efficient, minimizes contamination risk, and produces samples of intact diatoms. This method is designed to complete sample preparation within 24 h, sampling the bone for diatoms internally and externally. This method was developed using porcine long bones submerged in water with live diatoms for up to 3 months. Three marrow samples were extracted from each bone so the method was developed using 102 marrow samples. Additionally, 132 surficial bone and environmental samples were collected and prepared during method development. To briefly summarize the method, the bone joints were cut off with an angle grinder in a biosafety hood to expose the marrow, which was removed from the hip, knee, and shaft as separate samples. The marrow was digested with nitric acid at 400°C in glass beakers before being centrifuged with DI water, plated onto microscope slides, and observed with a compound microscope. Observation found good preservation of unbroken diatom cell walls throughout the process. This method can be used to prepare diatoms as forensic trace evidence.