Bacteria adhere strongly to the holocellulose
component characteristic of Sphagnum mosses, and this interaction
may form an important part of the remarkable preservative nature of peat bogs. |
|
Figure 1. The Tollundmannen is one of the
most famous proofs of the preservation efficiency of peat bogs. After being
buried in a Danish peat bog for 2200 years, the body was recoved remarkably
intact (Turner, R. C and Scaife, R. G. (1995). Bog Bodies - New Discoveries
and New Perspectives. London: British Museum Press.) Figure 2. Literature
Painter, T. J. (1983 a). Residues of d-lyxo-5-hexosulopyranuronic
acid in Sphagnum holocellulose, and
their role in cross-linking. Carbohydrate
Research 124, C18 - C21. Painter, T. J. (1983 b). Carbohydrate
origin of aquatic humus from peat. Carbohydrate
Research. 124, C22 - C26. Painter, T. J. (1991 a). Lindow Man,
Tollund Man and other peat-bog bodies: the preservative and antimicrobial
action of sphagnan, a reactive glycuronoglycan with tanning and sequestering
properties. Carbohydrate Polymers 15, 123 - 142. Painter, T. J. (1991 b). Preservation in
peat. Chemistry. & Industry.
(London), 421 - 424. Painter, T. J. (1998). Carbohydrate polymers
in food preservation: an integrated view of the Maillard reaction with
special reference to discoveries of preserved foods in Sphagnum - dominated peat bogs. Carbohydrate Polymers 36,
335-347. Painter, T. J. (2000). Preservation in Sphagnum – dominated peat bogs. In:
Briggs, D. E. G., Crowther, P. R. (Eds.). Palaeobiology
II. Oxford: Blackwell Science. Børsheim, K. Y., B. E. Christensen, T. J.
Painter (2001) Preservation of fish by embedment in Sphagnum moss, peat, or
holocellulose: experimental proof of the oxopolysaccharidic nature of the
preservative substance and its antimicrobial and tanning action. Innovative Food Science and Emerging
Technologies. 2, 63-74. |
K. Y. Børsheima,*, B. E. Christensenb,$
and T. J. Paintera,¶ aDepartment of Biotechnology and bNorwegian
Biopolymer Laboratory (NOBIPOL), Norwegian University of Science and
Technology, N-7491 Trondheim, Norway Introduction
Peat bogs formed of Sphagnum
mosses have remarkable capacity to preserve organic materials. Objects which
under other circumstances are easily mineralized by heterotrophic organisms, have been found to be preserved
in peat bogs. Archaeologists have collected documentation of this phenomenon,
including household utensils holding materials like butter, and various well
preserved bodies including human specimens. During a study of the
preservation processes that occur in peat bogs, we discovered an interaction
between the Sphagnum moss
polysaccharides and bacteria, that may play an important role in peat bog
preservation. We have elsewhere described other important preservative
effects, basically consisting of chemical reactions involving highly reactive
carbonyl groups in Sphagnum
holocellulose, binding ammonium and amino acids (Painter 1983a,b, Painter
2000). In this poster we suggest one more mechanism, which consists of direct
binding of bacterial cells to the Sphagnum
holocellulose. Peat bogs consists mainly of dead sphagnan plant material, which seems
to be close to resistant to biological degradation. Foreign organic material
incidently introduced into the interiour of peat bogs may also be preserved
for considerable amounts of time (Figure 1). Aims and objectives
To
test the effect of Sphagnum holocellulose on the concentration of various
types of bacteria in suspension.
Material
and Methods
Sphagnum holocellulose was prepared from leaves of peat moss (Sphagnum palustre). The leaves were
first washed with acetone to remove pigments etc., and hot extraction with 1%
sodium chlorite was used to remove lignin. This procedure leaves the
structural carbohydrate from the plant, which appears as a white, amorphous
substance which is almost insoluble in water. The structure of the
holocellulose is not yet completely resolved, but 5-keto-d-oxymannurose (5KMA) constitutes
approximately 20% of the molecule (Painter 1991a,b, Børsheim et al. 2001).
The pure holocellulose has been shown to act as a strong preservative
(Børsheim et al. 2001). In preservation experiments we used holocellulose
which had been reduced by borohydride as a control substance. This treatment
removes the reactive carbonyl groups on the 5KMA, which also turns off the
preservative effect. As a double
nonpreservative control we used wood cellulose. Various cultures of bacteria were
diluted in sterile buffer, and holocellulose was added. For practical reasons
the cellulose was enclosed in a kind of ”teabag” which was sewed from
plankton gauze of 60mm mesh size. Bacteria abundance was
measured before and after the addition of the cellulose using plate counts. Results
The pure Sphagnum holocellulose removed a very
large portion of the bacteria, regardless of bacterial type among the strains
tested (Table 1). Some removal of bacteria can be expected in porous
materials, but only a small part because the controls, both the borohydride
treated holocellulose and the wood cellulose, removed only a few percent of
the plate counts compared to the holocellulose. We have previously shown that the
carbonyl groups on the 5KMA residues in the holocellulose reacts with amino
groups, and binds covalently to a variety of primary amines including
proteins and pure ammonium (Painter 1998). Since proteins are abundant on
bacterial surfaces, it is not surprising that also bacterial cells are
immobilised by the holocellulose reactivity. The ability to arrest bacteria
irreversibly may form a part of the amazing preservative capacity of peat
bogs. Table 1. Immobilization
of bacteria with Sphagnum
holocellulose. Bacterial concentration in the treated culture after 20 hours
of incubation, compared the control (Borohydride reduced holocellulose) Bacterial culture % compared to the control Escherichia coli < 0.4
Bacillus sp. < 0.25
Micrococcus sp. <10 Local isolate* < 0.1 *The strain was
isolated from a preservation experiment. The preserving
properties of Sphagnum mosses are
probably mediated by several mechanisms acting in concert. We have previously
shown the effect of three separate mechanisms: ·
Exoenzymes
formed by microorganisms which are necessary to hydrolyse organic polymers
are immobilized and inactivated because they rapidly form Schiff’s bases with
the reactive carbonyl groups abundantly present in the Sphagnum holocellulose. ·
The Sphagnum holocellulose reacts with
organic material in general and the reaction products are inaccessible to
normal enzymatic degradation. These reactions are superficially the same as
the reactions taking place under industrial tanning of leather, and they lead
to similar products. ·
By forming
Schiff’s bases with free ammonium, bonded nitrogen is removed from the bog
water, consequently the microbes in the system experience severe nutrient
limitation. (references: Børsheim et al. 2001, Painter 1991a,b, Painter 1998) Conclusion
When we performed
experiments with pure cultures of bacteria, we found that all bacterial
species investigated adhered strongly to Sphagnum
moss, peat and pure holocellulose from Sphagnum
moss. The adhesion was strong, and occurred regardless of cell wall type such
as inferred from the Gram reaction. We are planning to investigate how the
strong adhesion of the bacterial cells to the plants influences the
metabolisms, viability and survival of the bacteria. Presently we are
confident that solid surfaces in natural peat bogs have a large capacity to
remove bacterial cells from the aqueous phase of the environment, by
reactions between highly reactive carbonyl groups in the Sphagnum holocellulose, and the bacterial surfaces. |