This article appeared in a journal published by Elsevier.

The attached
copy is furnished to the author for internal non-commercial research
and education use, including for instruction at the authors institution
and sharing with colleagues.
Other uses, including reproduction and distribution, or selling or
licensing copies, or posting to personal, institutional or third party
websites are prohibited.
In most cases authors are permitted to post their version of the
article (e.g. in Word or Tex form) to their personal website or
institutional repository. Authors requiring further information
regarding Elseviers archiving and manuscript policies are
encouraged to visit:
http://www.elsevier.com/copyright

Author's personal copy

Forensic Science International 184 (2009) 3741

Contents lists available at ScienceDirect

Forensic Science International

journal homepage: www.elsevier.com/locate/forsciint

Development and standardization of a microalgae test for determining deaths

by drowning
P.A. Daz-Palma a,*, A. Alucema a, G. Hayashida a, N.I. Maidana b
a
b

Department of Chemical and Pharmaceutical Sciences, Universidad Catolica del Norte, Antofagasta, Chile
Natural and Exact Sciences Faculty, Universidad de Buenos Aires, Buenos Aires, Argentina

A R T I C L E I N F O

A B S T R A C T

Article history:
Received 15 November 2007
Received in revised form 8 September 2008
Accepted 21 November 2008

The diatom test for the diagnosis of drowning is widely used in countries of the Northern Hemisphere
such as France and Japan. In Latin America, however, it has not been adopted as a routine procedure in
forensic autopsies. In aquatic ecosystems, dinoagellates and some chlorophytes are microalgae that,
like diatoms, have cell walls and other resistant structures. As a result, they can be found in tissues from
drowning victims, which is important because diatoms may be rare under particular environmental
conditions. On this basis, we propose to extend the diatom test to include other microalgae for the
determination of death by drowning.
In this work, we developed a standardized procedure for detecting microalgae in tissues from
drowning victims, with techniques described in the international literature and designed by us. The
corpses were recovered from coastal areas in Antofagasta Region, Chile, during summer 2005. The most
effective procedure for the treatment of water and tissue samples involved the combination of enzymatic
digestion (proteinase K) and chemical digestion. The technique allowed the recovery of dinoagellate
evidence belonging to genera Prorocentrum, Ceratium, Dinophysis and Protoperidinium; silicoagellates of
the genus Dictyocha; an undetermined, lamentous chlorophyte; entire valves of centric diatoms and
fragments of pennate and centric diatoms. This is the rst protocol using microalgae other than diatoms
for forensic cases in Latin America, and particularly in Chile.
2008 Elsevier Ireland Ltd. All rights reserved.

Keywords:
Drowning
Microalgae
Diatoms

1. Introduction
Medicolegal investigation relied largely on laboratory-based
analyses, and some of these, like the DNA prole of body uid, have
become valuable routine methods [1,2]. The so-called diatom
test for the diagnosis of death by drowning is widely practiced in
Northern Hemisphere countries like France and Japan, but is not
being used regularly in forensic practice in Latin America.
Microalgae are of limited application in forensic sciences, but
they play an important role in certain types of research [1,2], thus
emphasizing the necessity of studies on their taxonomy and
ecology. In fact, evidence provided by taxonomists and ecologists
can be presented by either the plaintiff or defendant to support a
verdict [1,2].
In forensic science, detection of diatoms in tissues may
contribute to the diagnosis of drowning, which is a very common

* Corresponding author at: Department of Chemical and Pharmaceutical

Sciences, Universidad Catolica del Norte, Avenida Angamos 0610, Antofagasta,
Chile. Tel.: +56 55 355624/5383; fax: +56 55 355632.
E-mail address: pdiaz@ucn.cl (P.A. Daz-Palma).
0379-0738/$ see front matter 2008 Elsevier Ireland Ltd. All rights reserved.
doi:10.1016/j.forsciint.2008.11.015

cause of death. The circumstances can be established by eyewitnesses testimony or a suicide note, but when underlying causes
are unclear, the standardization of methodologies helps to
determine whether a crime was committed [1]. Death by drowning
is easily recognized in a fresh cadaver, but histopathological signs
are hampered by the deleterious effects of decomposition, or by
strong waves and currents crashing the body against the rocks.
Therefore, in case of severe injuries, the actual cause of death
should be ascertained [1].
The diatom test has been accepted by many authors [39], but
there is no consensus on whether diatoms are accurate indicators
of drowning since they have been also found in non-drowning
victims, such as swimmers and shermen [10,13]. These ndings
led some researchers, for example Gylseth and Mowe [11],
Shellman and Sperl [12] and Foged [14], to rule them out as
reliable evidence.
Nevertheless, it is reasonable to assume that such cases may
represent false positives resulting from sample contamination
prior to the analysis.
On the other hand, Peabody [7,8] and Auer and Mottonen [15]
state that diatom count can be used to discriminate between
drowning and non-drowning cases.

Author's personal copy

P.A. Daz-Palma et al. / Forensic Science International 184 (2009) 3741

38

Table 1
Results obtained with the different digestion techniques.
Procedure no.

Concentration

Digestion time (h)

Results

1 (HCl)

5%
10%
20%
50%
80%
Concentrated

2 and 24
2 and 24
2 and 24
2
2
2

Partial digestiona, there is still a large amount of organic matter.

5%
10%
50%
80%
Concentrated

2 and 24
2 and 24
2
2
2

Partial digestion, there is still a large amount of organic matter.

3 (H2O2)

20%
Concentrated

2
2 and 24

Partial digestion, there is still large amounts of organic matter.

4 (combined)

conc. H2O2, and HCl 20%

0.05 +

Optimum digestionc, there is a low amount of organic matter and

the number of species recovered is low.

2 (H2SO4)

a
b
c

Aggressive digestionb, there is a decrease in the amount of organic

matter but the number and diversity of species recovered are low.

Aggressive digestion, there is a decrease in the amount of organic matter,

but the number and diversity of species recovered are low.

The amount of organic matter is higher enough to hinder the visualization of microalgal structures.
Destruction of microalgae frustules and thecae.
The removal of a larger amount of organic matter from the sample allows the clear visualization of the microalgal structures.

Taking into account the small number of diatom valves that can
be recovered from cadaveric tissue [15], a test using microalgae
other than diatoms (e.g. dinoagellates and chlorophytes) may
provide a more reliable diagnosis.
It is worthwhile to mention that both the diatom test and the
microalgae test proposed by us do not constitute by themselves a
conclusive proof of death by drowning, from a legal point of view.
Instead, they represent tools that, together with other analyses
(toxicology testing, determination of chloride and strontium levels
in blood and presence of hemoglobinemia, etc.), help the Public
Prosecutors to reach a nal decision.
Diatoms and other algae that can be of use in forensic
investigation are identied by the morphology of the frustule,
theca, endoskeleton, cell wall, etc. The observation of these
structures requires the removal of organic matter represented by
microalgae content and tissue debris. The three most common
methods used for preparing samples for diatom analysis involve
treatment with a strong acid, tissue solubilizers and/or enzymatic
digestion [16]. However, some of these methods fail when used for
marine diatoms [16] and consequently there is an increasing need
for developing methodologies that can be extrapolated to different
aquatic systems.
In the present work we present a standardized procedure for
diagnosing death by drowning. In addition, we compare different
digestion techniques to determine which provides the maximum
amount of forensic evidence (microalgae) for taxonomical
identication. The techniques were performed using bone marrow
and water samples, and were designed by us or obtained from
literature.

2. Methods
All plasticware used was carefully washed with free-phosphorus detergent and
soaked in 20% HCl for 24 h, rinsed with distilled water and nally rinsed with
diatom-free distilled water (DFDW), obtained by ltration through a 0.45-mmpore-size nitrocellulose membrane.
Tissue samples were obtained from autopsies performed in the Medicolegal
Service of Antofagasta City (Chile). The samples came from three individuals drown
during summer 2005; one neonate, who died of sudden death that same year,
served as control. A sample of sternal bone marrow was taken from each individual,
and a lung sample from one of them. Each sample was washed with DFDW before
cooling at 4 8C.
Almost simultaneously with the nding of the victims, seawater samples were
collected along the coast of Antofagasta Bay, close to the sites where bodies were
recovered. Samples were taken in triplicate with a 45-mm mesh plankton net, xed
in 2% formalin and stored at 4 8C until processing.

Prior to any treatment, seawater samples were examined under the microscope
to estimate qualitatively the density of microalgae, and they were concentrated by
evaporation and centrifugation when the number of microalgae was small.
We chose as the best digestion method the one yielding the cleanest sample, the
largest number of evidences (complete valves, thecae and endoskeletons, or
identiable microalgal fragments), and the optimum visualization of the characters
for species identication. To determine the most appropriate procedure, we
modied methods widely reported in the literature by varying digestion time and
type and concentration of reagents. We also tested an alternative method
(Treatment 4) applied by one of the authors to another type of diatom analysis [17].
Bone marrow and water samples were treated as follows (see Table 1 for
details)Treatments 1 and 2, digestion with strong acids HCl and H2SO4 [5]: acid
concentration were 5 and 100%, and digestion times were 2 and 24 h, respectively.
Treatment 3 (enzymatic): digestion using a solution of 0.5 U/ml proteinase K and
0.01 M TrisHCl, pH 7.5, with 2% sodium dodecyl sulfate (SDS), neutralized with
concentrated HCl [6].
Treatment 4: 12 ml of concentrated hydrogen peroxide (H2O2) were added to
each sample. The samples were put in a double boiler and heated in the microwave
at 80% power (around 650 W) for 3 min. After cooling, 1 to 2 ml of 20% HCl were
added. Following Treatments 1, 2 and 4, samples were neutralized by successive
washes with DFDW and centrifugations.
The treated water samples were observed in permanent preparations mounted
on a hydrophobic adhesive medium (Entellan or Eukitt), whereas tissue samples
were observed in temporary preparations. Samples were examined under a
binocular microscope Zeiss Axiostar equipped with a digital camera System
Olympus DP70.
On the basis that bone marrow samples were taken from subjects known to have
died by drowning, we veried the efciency of the procedure proposed in this work,
by comparing the number of complete and incomplete diatoms obtained from
tissues with reference values given by Ludes and Coste [1].
We compared the species composition of microalgae in seawater and bone
marrow samples with the Jaccard index of similarity [18], using the BioDiversity
PRO program (version 2).

3. Results
The results of the chemical digestion techniques are shown in
Table 1. We selected Treatment 4 for water samples because it
achieved the best performance.
The enzymatic Treatment 3 resulted in partial digestion of bone
marrow and lung samples and therefore microalgae observation
was hindered by the large proportion of organic matter (Fig. 1).
However, results improved when Treatment 4 was applied after
Treatment 3. The combination of the enzymatic and chemical
digestions increased the number of the specimens recovered, with
most of the microalgal structures (frustules, thecae and endoskeletons) being complete. This allowed the identication of a high
diversity of taxa including diatoms, dinoagellates, silicoagellates
and chlorophytes, thus increasing the accuracy of the forensic
diagnosis.

Author's personal copy

P.A. Daz-Palma et al. / Forensic Science International 184 (2009) 3741

Fig. 1. Map of the Baha de Antofagasta, Chile, showing the location of the sites
where victims of drowning were recovered and, where seawater samples were
collected.

Table 2
Results obtained with the bone narrow sample from individuals: no. of complete
frustules, thecae or endoskeletons/no. of incomplete frustules, thecae or
endoskeletons/total of algal remains.
Class

Individual 1

Individual 3

Diatoms (Bacillariophyceae)

Silicoagellates (Dictyochophyceae)
Green algae (Chlorophyceae)

2/0/2
1/3/4
0/3/3

183/0/183
1/1/2
8/10/18
22/21/43
1/0/1

0/3/3
1/0/1
2/0/2

0/4/4

Total

217/38/256

3/7/10

Dinoagellates (Dinophyceae)

Table 3
Number of expected valves per mass unit (*) and observed valves for samples 2 and
4. (*) According to Ludes and Coste [12].
Sample No.

Sample weight (g)

Expected valves

Observed valves

1
3

3.52
5.68

1.76 2*
2.84 3*

9
4

39

The control sample was negative for microalgae, and showed fat
cells some blood cells like megakaryocytes and bone structures like
haversian canals.
In the sample from subject 1 (Table 2), we identied taxa of
diatoms, dinoagellates and silicoagellates. The number of
dinoagellates and silicoagellates was comparable to that of
diatoms. With regard to diatoms, the proportion of complete
valves was higher than that of incomplete ones for most of the taxa
(Table 2). No marine microalgae were recovered from the lung
sample of subject 2.
Table 2 also shows the taxonomic diversity found in the sample
of subject 3. The sample contained different taxa, including
diatoms, dinoagellates and silicoagellates. Microalgal structures
were less abundant than in the sample from subject 1 and also
there were fewer structures for each taxon (Table 2). Incomplete
valves, thecaes and endoskeletons outnumbered complete ones. As
in the sample from subject 1, the silicoagellates, represented
uniquely by Prorocentrum micans, were the only ones presenting
complete thecae.
Table 3 shows that the counts of diatom valves in the samples
from subjects 1 and 3, expressed per unit of tissue, were higher
than those established by Ludes and Costes [1]; this was
particularly the case for the sample from subject 3, who was
found in the warm waters of a small semi-closed bay with
microalgae blooms occurring almost all-year-round at intervals
only depending upon the tidal regime (Gob. Martima de
Antofagasta, personal communication).
Table 4 shows the microalgae identied in water samples taken
at the coasts of beaches El Trocadero, Los Pinares and Juan Lopez.
The water samples from El Trocadero and Los Pinares were
analyzed together with the tissue sample of subject 1 because the
corpse was found in the midway between the two beaches. We
identied seven and six different taxa at El Trocadero and Los
Pinares, respectively, with diatoms, dinoagellates and silicoagellates represented at both sites.
The microalgae from the water sample taken at the coast of the
locality Juan Lopez (Table 4); was compared with the tissue sample
of subject 3, who was recovered at this same site. We identied 15
taxa including diatoms, dinoagellates and silicoagellates.
Dinoagellates showed the highest abundance of microalgal
structures and species richness in both water and tissues samples;
in particular, the one responsible of the red tides occurring in the
region during summer [19].
The values of the Jaccard index of similarity obtained for water
and bone marrow samples are shown in Table 5. Hurlimann et al.
[15] stated that algae from water and tissue samples belong to the
same community when the similarity index is higher than 60%.
According to this, there was a low similarity between bone marrow

Table 4
Algal remains recovered in the samples from different localities. FTC, number of complete frustules, thecae or endoskeleton; FTI, number of incomplete frustules, thecae or
endoskeleton; R, replicate; x, mean values; S.D., standard deviations. Localities ET: El Trocadero; LP: Los Pinares and JL: Juan Lopez.
R1

Locality

Total

FTC

FTI

Total

FTC

FTI

Total

Diatoms

ET
LP
JL

5
4
42

14
4
33

19
8
75

14
11
141

24
0
32

38
11
173

0
5
47

0
10
19

0
15
66

28.5
11.4
104.5

26.1
4.7
78.4

Dinoagellates

ET
LP
JL

101
35
30

22
1
96

123
36
126

254
29
24

1
3
0

255
32
24

0
13
16

0
2
7

0
15
23

189
27.6
57.7

117.4
16.1
62.3

Total locality

ET

106

36

142

268

25

293

217.5

143.6

Total locality

LP

39

44

40

43

18

12

30

39

Total locality

JL

72

122

201

165

32

197

63

26

89

162.2

FTC

R2

Algal remains

FTI

R3

S.D.

20.7
140.6

Author's personal copy

P.A. Daz-Palma et al. / Forensic Science International 184 (2009) 3741

40

Table 5
Jaccard indices of similarity (JIS%) between bone marrow and water samples.
Locality

Bone marrow samples

Sample 2

Los Pinares
El Trocadero
Juan Lopez

Sample 4

55.55
50.00
28.57

samples and seawater samples taken from the site where bodies
were found because indices ranged between 29 and 55%.
4. Discussion
The enzymatic digestion treatment was more effective to
remove the organic matter from human tissues than from
microalgae. On the other hand, the addition of sodium dodecyl
sulfate detergent (SDS) was found to be effective in causing
membrane lysis and protein degradation, as well as in reducing
lipids.
In the chemical digestion treatment, the addition of hydrogen
peroxide at high temperature was effective in removing organic
material. Hydrogen peroxide is a very powerful oxidant, which is
decomposed into products at different oxidation states under high
temperature conditions, since it oxidizes and reduces itself.
In most cases, the digestion using strong acids was less
aggressive due to the low concentration of chloride acid (20% HCl)
and shortening of the process. This was reected in the
experiments performed to determine the most appropriate
chemical digestion procedure. Thus, at a concentration of 20%
HCl, the number of taxa was smaller at longer digestion times.
Likewise, a similar result was obtained at concentrations above
20%.
Finally, the combination of enzymatic and chemical digestions
in tissue samples resulted in a synergism that substantially
increased the diversity of diatom, dinoagellate and chlorophyte
species. Consequently, we conclude that this combination yields a
better performance than other procedures reported in literature
[1,20,21].
Silica is scarcer in marine than in freshwater environments,
thus representing a limiting factor for seawater diatoms. These
compete with other species that also incorporate this element, and
their valves contain less silica than freshwater diatoms [22]. As a
result, seawater diatoms are vulnerable to aggressive chemical
treatments, and this accounts for the efciency of the combined
digestion method used here. This interpretation also applies to
silicoagellates because of their siliceous endoskeletons [23].
Dinoagellate thecae, which are composed of glucan [23], were
resistant to the combined chemical and enzymatic treatment. The
nding of a chlorophyte in bone marrow samples (Table 2) was
surprising because its external structures are usually vulnerable to
strong acids, and supports our conclusion that this technique is
successful in recovering a large number of taxa.
Despite the fact that both chloride and sulfuric acids are
classied as strong acids, the former was less aggressive than the
latter based on the larger number of evidences obtained from each
sample.
An appropriate digestion of the organic matter from intra- and
extra-cellular components is needed to improve the visualization
of structures used for microalgal identication.
We obtained a higher diversity of taxa (diatoms, dinoagellates,
silicoagellates and chlorophytes) from tissue samples than that
reported in other studies [14,24]. This result indicates that,
conversely to the traditional methods, the combined procedure
allows the recovery of a larger number of evidence and does not

depend exclusively on habitat features conditioning the availability of diatoms.

The lung is expected to show the highest diversity of taxa
because it is one of the rst organs to come into contact with water
during the drowning process, and from which water enters the
bloodstream to reach the different tissues. This discordance with
our results is likely to be due to the small sample volume, and
reects the importance of using samples from different sources
(bone marrow, lung, blood) of the same subject to validate the
procedure.
A small number of non-marine diatom valves were recovered
from the lung sample; these are frequently found in local
continental environments [17]. Their presence can be attributed
to contamination occurring while the victim was alive because the
valves, which can be easily transported by the wind, may have
entered into the lung.
In addition, contamination could have taken place during
corpse manipulation, autopsy, laboratory analysis, or in any
other stage of the process. The efcacy of the method used in the
present work is supported by the large number of valves and
fragments recovered from tissue samples of subjects 1, 2 and 3,
which even exceeded the minimum number suggested by Ludes
and Coste [1]. With regard to algae other than diatoms that can
be recovered by this technique, we propose to standardize the
minimum number of structures required to consider the test as
positive.
According to the Jaccard similarity indices, algae found in bone
marrow and water samples were not likely to belong to the same
community. This result is probably due to environmental factors
not considered in our analysis on the basis that water samples
were not taken simultaneously with the nding of the body.
The probability of microalgae to reach tissues during submersion may depend on their morphology. Thus, the maximum lengths
of several taxa exclusively found at the site of drowning were
higher than the average length of algae from the tissue, such as
Synedrosphenia sp. and Fragilaria fasciculata. In this respect, studies
on the use of different morphological features as forensic evidence
are needed to validate the Jaccard index of similarity of 60%
proposed by Hurlimann et al. [15].
5. Conclusion
We developed a reproducible protocol for both water and tissue
samples. The digestion procedure used in the present study
increased the recovery of evidences (microalgae taxa) in comparison with various methods usually referred to in literature. In
addition this novel technique facilitates the visualization of
representative microalgal structures and provides a number of
undamaged cell coverings large enough to allow an accurate
taxonomic identication. The importance of this result lies in the
fact that traditional techniques are very aggressive and cause not
only the digestion of tissues, but also the fracture of most of the
algal material.
We established that microalgae found in one of the analyzed
tissue samples did not enter during drowning based on the
detection of foreign material, which emphasizes the importance of
identifying local microalgal ora.
A standardized microalgal test for the diagnosis of death by
drowning provides valuable evidence for the interpretation of
forensic cases. However, this evidence must be contrasted with
that obtained from other sources because of the numerous factors
involved in the accident, such as the type of environment where
drowning took place (marine or freshwater ecosystems), the life
history of victims, presence of eyewitnesses, and routine
procedures performed by qualied forensic professionals (e.g.
toxicologists).

Author's personal copy

P.A. Daz-Palma et al. / Forensic Science International 184 (2009) 3741

Our results contribute to enhance the current knowledge of

forensic biology, and the microalgal test proposed here appears as
a valuable, reliable and sensitive diagnostic tool to be used in
routine forensic procedures.
Acknowledgments
We thank Drs. Carlos Contreras O. and Jaime Llanos S. for critical
reading of the manuscript. Other acknowledgements to Miss
aloza for the analysis of diatoms samples.
Karina Pen
References
[1] B. Ludes, M. Coste, Diatomees et medicine legal, Med. Int., Francia, 1996.
[2] M.S. Pollanen, Diatoms and homicide, Forensic Sci. Int. 91 (1) (1998) 2934.
[3] N.I. Hendey, The diagnostic value of diatoms in cases of drowning, Med. Sci. Law
13 (1973) 2334.
[4] D.A. Neidhart, R.M. Greedyke, The signicance of diatom demonstration in the
diagnosis of death by drowning, Am. J. Clin. Pathol. 48 (1967) 377382.
[5] J.V. Pachar, J.M. Cameron, Submersion cases: a retrospective study 19881990,
Med. Sci. Law 32 (1992) 1517.
[6] A.J. Peabody, Diatoms in forensic science, J. Forensic Sci. Soc. 17 (1997) 8187.
[7] A.J. Peabody, Diatoms and drowning: a review, Med. Sci. Law 20 (1989) 254261.
[8] J. Timpermann, The diagnosis of drowning. A review, Forensic Sci. 1 (1962) 397
407.
[9] K. Krammer, H. Lange-Bertalot, Susswasserora von Mitteleuropa. Bacillariophyceae 1, 2, 3 and 4, G. Fisher, Jena, 19881991.

41

[10] M. Kobayashi, Y. Yamada, W. Zhang, Y. Itakura, M. Nagao, T. Takatori, Novel

detection of plankton from lung tissue by enzymatic digestion method, Forensic
Sci. Int. 60 (1993) 8190.
[11] B. Gylseth, G. Mowe, Diatoms in lung tissue, Lancet 29 (1979) 1375.
[12] B. Schellman, W. Sperl, Nachweis im knochenmark (Femur) nichter-trunkener,
Rechtsmed 83 (1979) 319324.
[13] N.I. Hendey, Diatoms and drowning, Med. Sci. Law 20 (1980) 289.
[14] N. Foged, Diatoms and drowning, Forensic Sci. Int. 21 (1983) 153159.
[15] A. Auer, M. Mottonen, Diatoms and drowning, Rechtsmed 101 (1988) 8798.
[16] J. Hurlimann, P. Feer, F. Elber, R. Niederberger, R. Dirnhofer, D. Wyler, Diatom
detection in the diagnosis of death by drowning, Int J. Legal Med. 114 (2000) 614.
[17] L. Sidari, N. Di Nunno, F. Costantinides, M. Melato, Diatom test with Soluene-350
to diagnose drowning in sea water, Forensic Sci. Int. 103 (1999) 6165.
[18] C. Daz, N.I. Maidana, Diatomeas de los salares Atacama y Punta Negra, II Region
Chile, Centro de Ecologa Aplicada Ltda. y Minera Escondida Ltda. Santiago, Chile,
2005.
[19] P. Jaccard, Etude comparative de la distribution orale dans une portion des Alpes
et du Jura, Bull. Soc. Vaud. Sci. Nat. 37 (1901) 547579.
[20] L. Rodrguez, Observaciones sobre efectos de El Nino 19971998 en el toplancton de baha Antofagasta y durante un fenomeno de surgencia en baha Mejillones
del Sur, Antofagasta, Chile, CONA (2004) 97118.
[21] B. Ludes, M. Coste, M. Tracqui, P. Mangin, Continuous river monitoring of the
diatoms in the diagnosis of drowning, J. Forensic Sci. 41 (1996) 425428.
[22] B. Ludes, S. Quantin, M. Coste, P. Mangin, Application of a simple enzymatic
digestion method for diatom detection in the diagnosis of drowning in putreed
corpses by diatom analysis, Int. J. Legal Med. 107 (1994) 3741.
[23] F. Thomas, W. Van Hecke, T. Ghent, The medicolegal diagnosis of death by
drowning, J. Forensic Sci. 8 (1962) 18.
[24] O. Parra, M. Gonzalez, V. Dellarossa, Manual taxonomico del toplancton de aguas
continentales: I Cyanophyceae, III Cryptophyceae-Dinophyceae-Euglenophyceae,
V Clorophyceae 1 y 2, Universidad de Concepcion, 19821983.