Microalgae Test For Determining Death By Drowning

DEVELOPMENT AND STANDARDIZATION OF A MICROALGAE TEST FOR DETERMINING DEATHS BY DROWNING By Sanya-Isijola, Ademuyiwa 36641 INTRODUCTION Deaths that occur as a result of drowning are often attributed to accidents, suicide or murder. Knowing the exact cause that led to the drowning of the victim is down to forensic investigations conducted. “The world incidence of death by drowning is estimated at about 5.6% per 100,000 of population. Approximately 1,500 deaths from drowning occur in the UK each year, 25% occur in the sea and the rest in inland waters; majority of the victims are young adults and children; two-third are accidental and one-third are suicidal, homicide by drowning is very rare”-from Bodies from water, Dept of Forensic medicine, University of Dundee. When corpse is found in water, the cause of death is often associated with drowning, the body recovered from water prompts two questions:  

Was the victim alive or dead when he entered the water? Is the cause of death really drowning? If not, what is the cause of death?

To answer the questions listed above, the following information must be available   

The circumstances preceding the death The circumstances of recovery of the body The autopsy findings [1]

What is Drowning? Drowning occurs when there is lack of oxygen reaching the body tissues due to immersion in water. This is called ASPHYXIA. [2] Asphyxia occurs when there is an interference with the mechanics of breathing, thus affecting regular respiratory functions. There are several classifications for Asphyxia which include Suffocation, Strangulation, Hanging, drowning etc. Types of Drowning There are different types of drowning, they are:

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Fresh water drowning

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Salt water drowning

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Dry drowning [3]

Death by drowning primarily occurs when certain areas of the body e.g tissue are deprived of adequate oxygen supply (Hypoxia), thus excluding air from the lungs. It can also be as a result of electrolyte imbalance or/and Haemolysis which occurs when water is inhaled by affecting the blood. When water is inhaled into the lungs, fine froth is produced and found lining the Trachea as a result of the mixture of the water and mucus due to respiratory movement.

Autopsy findings The following are some of the possibly identified findings that are discovered by Pathologists in the course of autopsy carried out on corpse taken from water. External findings 

Roughening of the skin (Anserina cutis)

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Skin swelling and wrinkling (Skin Maceration)

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Signs of suggestive drowning i.e Presence of objects (weed, sand) clutched in the hands of the victim is an indication that victim was alive before getting into the water.

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Transformation of the fatty layer beneath the skin into soap like material. This process (Adipocere) takes weeks to occur and prevents the decomposition of the corpse in water.

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Presence of fine white froth in the mouth and nostrils

Internal Findings 

Inflated and sodden lungs.

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Presence of fine froth lining the Trachea, Bronchi and distal air passages.

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Presence of water in the stomach, this is true in 70% of the cases.

Diagnosis difficulties When drowning occurs, the lungs are over inflated and heavy with fluid, this is not enough to conclude that death was caused by drowning because death associated with pulmonary edema has a similar effect on the lungs. Pulmonary edema is present when death occurred as a result of drug over dose or heart failure. Pulmonary edema is fluid accumulation in the lungs. It leads to impaired gas exchange and may cause respiratory failure. (Wikipedia) It is difficult to obtain pathological proof to show that death occurred by drowning because of the similarities found in the trachea (froth) and lungs of the corpse from water and the lungs of victim whose death occurred as a result of diseases (Epilepsy, heart disease), alcohol or drug over dose or cardiac arrest. Even after bodies are found immersed in fresh or salt water, in some cases, it is still difficult to say whether death was due to drowning or not, this is a major concern for the Pathologist. In order to ascertain the cause of death, other signs that may have caused death are thoroughly investigated and excluded one after the other. If circumstances of death are known, the diagnosis of drowning will be less difficult. Hence, diagnosis of drowning is made on the basis of exclusion by doing the following:    

Complete autopsy Toxicological screening Histologic analysis of all organs (lungs inclusive) Diatom test [4]

DIATOM IDENTIFICATION The diatom test is yet to be a regular practice in latin America but has been widely used in countries like France and Japan. Diatom analysis involves using the following methods on tissues: 

Strong acid

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Tissue solubilizers

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Enzymatic digestions

These methods are known to fail when used for marine diatoms, hence the need to develop different ways that can be used on different aquatic system.

Diatoms are microscopic, unicellular algae with heat and acid resistant shells called Frustules found in most fresh or sea water. The Frustule is made up of hydrated Silica dioxide; it also contains carbon and protein. [3] There are 15,000 species of Diatom known, approximately half in fresh water and the other half in sea water. In 1941, Incze demonstrated that during drowning, diatoms could enter the systemic circulation via the lungs. Diatoms are used for diagnostic test for drowning based on the hypothesis that unless victim was alive in water (i.e circulation is functional), diatoms will enter the systemic circulation and be deposited in organs like the bone marrow, kidney and the brain. The lung tissue is exempted during test because Diatom might be present in lungs of victim that was dead on getting to water. During autopsy, an acid digestion of these tissues is carried out and confirming the presence of diatoms provides a strong evidence of drowning.

Environmental conditions that affect the potency of Diatom Test Due to certain environmental conditions, Diatoms may become rare in fresh water or sea water and may also be present in the tissue of victim on getting to water. Such conditions will affect the potency and accuracy of the Diatom test as prove of drowning, hence the need to develop and standardize the Microalgae test (Dinoflagellates and chlorophytes) for determining deaths by drowning. These environmental conditions include: 

The difficulty of excluding contamination: with the aid of contaminated foods consumed such as Salads and Shell fish ,diatoms may enter the circulation via the gastro-intestinal tract

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Diatoms are also present in small numbers in the air, they are found in the environment such as building industry and on dusting powder for rubber gloves. Hence, they may enter the circulation via the respiratory tract. [1]

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Environmental conditions such as seasons of the year (winter, summer etc), temperature, salinity and depth of water affect the presence of diatoms by either making them rare or abundant.

A research titled “Microscopic marine organisms in hydrography” done by Prof P.T Cleve of the University of Upsala, Sweden illustrated how the above mentioned environmental conditions (temperature, salinity etc) affect the population and presence of different types of Planktons/diatoms.

“There can be no doubt about the close connection between the state of the sea and movement of the air and the still obscure causes of the migration of fishes may be found to be intimately connected with the change in water containing different kinds of Plankton”. Prof P.T Cleve [5]

Standardized Microalgae Test An experiment was done by Daiz-Palma PA, Alucema A, Hayashida G and Maidana NI of the Chemical and Pharmaceutical science departments, Universidad Catolica del Norte, Chile to develop and standardize Microalgae test for determining deaths by drowning. The following were done in the cause of the experiment: 

Different digestion techniques were compared to ascertain which provides the highest amount of microalgae using the bone marrow and water samples.

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Tissue samples (lung and sterna bone marrow) were obtained from 3 different drowned victims.

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Sea water samples close to where the victims were found was also collected.

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The best digestion method was chosen based on the one yielding the cleanest sample.

Results 

It was noted that the enzymatic digestion treatment was more effective to remove the organic matter from human tissues than from the microalgae.

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When the enzymatic and chemical digestion was combined for use on tissue samples, it yielded better performance and increased the diversity of diatom, dinoflagellates and chlorophytes species.

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It was noted that chemical digestion was not favorable for sea water diatoms. Silica is scarcer in marine than fresh water environment (sea water diatoms contain less Silica than fresh water diatoms). Hence, combined digestion is used for sea water diatoms and Silico flagellates.

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It was noted that Dinoflagellate thecae was resistant to a combination of chemical and enzymatic treatment. [6]

Conclusion

The combined digestion procedure when used increased the recovery of a larger number of evidence (Microalgae) in comparison with various methods. This technique preserves the algae material (protects cell covering) that easily becomes fractured when the other aggressive traditional techniques are used. Hence, a standardized microalgae test provides valuable evidence that can be used in court. However, diatoms and microalgae tests are not conclusive proof of death by drowning, they are only additional tools used with other forms of diagnosis such as Toxicological screening, complete autopsy, histologic analysis of all organs, history of victims, eye witness presence etc on the basis of exclusion to ascertain the likely cause of death and reach a final decision.

Reference 

http://www.dundee.ac.uk/forensicmedicine/notes/water.pdf [1]

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http://users.ox.ac.uk/~oulsc/theory/drown.html [2]

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www.mona.uwi.edu/pathology/staff/escoffery/documents/Forensic%20II%20%20%20Asphyxia,%202007.DOC [3]

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http://netk.net.au/Reports/DiagnosticsOfDrowning.asp [4]

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http://sabella.mba.ac.uk/174/01/Microscopic_marine_organisms_in_the_service_of_hydr ography.pdf [5]

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Development and standardization of microalgae test for determining deaths by drowning by Daiz-Palma PA, Alucema A, Hayashida G and Maidana NI of the Chemical and Pharmaceutical science departments, Universidad Catolica del Norte, Chile [6]