Kartchner Caverns Microbial Observatory
National Science Foundation
photo of caverns

Research

The Impact of Tourism on Kartchner

Impact of Tourism on Kartchner

Nonculture-Based Analysis

Culture-Based Analysis

Fungi

Nematodes

Kartchner Caverns in Benson, AZ was opened for tourism in 1999 following a careful development protocol that was designed to maintain predevelopment conditions.  As a part of an ongoing effort to determine the impact of humans on this limestone cave, samples were collected from cave rock surfaces along the cave trail traveled daily by tour groups (200,000 visitors yr-1) and compared to samples taken from areas designated as having medium (30-40 visitors yr-1) and low (2-3 visitors yr-1) levels of human exposure. The Kartchner Cavern human impact study sites are shown below.

Kartchner Caverns Study Sites

Samples were also taken from painted fiberglass moldings installed during cave development to study a slime formation problem that was discovered on painted fiberglass surfaces within the developed portion of the cave. These molded fiberglass blocks and fittings were mounted to disguise plumbing and electrical wiring as well as maintenance areas that might detract from the visitors’ visual experience. In an effort to eliminate the slimy growths, the surfaces were washed with a 10% bleach solution.  However, the biofilm growths returned within one month of treatment.  Of primary concern, was the fact that bacteria potentially introduced into the cave following the installation of synthetic surfaces or left behind from the skin, clothing and shoes of people touring the cave might alter natural microbial communities found in the cave.  Thus, the objective of this initial study was to determine the source of the observed slime and to compare the culturable heterotrophic populations isolated from the fiberglass surfaces to those present on adjacent natural rock surfaces.


Culture Diversity in Kartchner

Culturable bacteria were recovered from samples taken from the high, medium, and low impact areas of cave as well as from fiberglass surfaces.  A total of 90 unique isolates were identified using 16S rRNA PCR and sequencing (Figure 1).

Figure 1. Phylogenetic tree showing the alignment of the 90 cultured 16S rRNA sequences from Kartchner Caverns.  This analysis resulted in a 1532-character dataset of which 757 characters (49.4 %) were variable and 691 characters (45.1 %) were parsimony informative.  Maximum-parsimony analysis of the 16S dataset yielded 3049 equally most-parsimonious trees (steps = 4482, CI = 0.326, RI = 0.788), which differed primarily in changes in relationships among taxa in terminal clades.  Five principle clades were evident in all trees: Firmicutes, Actinobacteria, α-Proteobacteria, β-Proteobacteria, and γ-Proteobacteria.  All principle clades were supported by bootstrap values >80%.  Three isolates were identified as Bacteriodetes and composed the root of the tree. Analysis of the truncated datasets (ambiguous section removal) had only minor effects on the resulting tree topology compared to that resulting from analysis of the complete dataset, and the position and composition of major clades and high bootstrap support remained unchanged (data not shown). 

Figure 1

Diversity generally decreased as human impact increased leading to the isolation of 32, 27, and 22 strains from the low, medium, and high impact areas, respectively. The degree of human impact was also reflected in the phylogeny of the isolates recovered.  Although most isolates fell into one of three phyla: Actinobacteria, Firmicutes, or Proteobacteria, the Proteobacteria were most abundant along the cave trail (77% of the isolates) while Firmicutes predominated in the low (66%) and medium (52%) impact areas (Figure 2).

Figure 2. Phylogenetic distribution of the 90 cultured isolates from high, medium, and low
impact sites as well as painted fiberglass surface

Figure 2. Phylognetic distribution.

While the abundance of Proteobacteria along the cave trail seems to include microbes of environmental rather than of anthropogenic origin, we hypothesize that their presence is a consequence of increased organic matter availability due to lint and other organics brought in by cave visitors.  This hypothesis is supported by a qualitative examination of the cave carbon gradient:

Pre-development there were two organic carbon sources in Kartchner: 

1.  DOC and colloidal organic matter in water from

      
2. Complex organic matter from

Post-development there is one additional carbon source – humans

Pre-development qualitative carbon analysis:

Rotundra-Throne area (high impact) had the lowest organic input

--No bats roost in this area
--The ceiling is too thick for root penetration
--The area is not subject to flooding
--Organic matter is introduced solely through infiltration

Grand Central Station (moderate impact) had highest organic input

--Bats roost in area
--The site is relatively shallow and close to the entrance
--The area is subject to minor floodingorganic matter is introduced from bat guano, cricket guano, and from infiltration and minor flooding

Subway Tunnel (low impact) had intermediate organic input

--No bats roost in this area, no root penetration
--The site has almost constant percolation of water
--The area has the highest amount of flooding
--Organic matter is introduced infiltration and flooding


Qualitative comparison of organic carbon pre- and post-development:

 
Pre-development
 
Post-development
High impact 
lowest
arrow
highest
Medium impact   
highest
arrow
intermediate
Low impact   
intermediate
arrow
lowest

Research outcome:  misters have been placed strategically in portal tunnels so that tourists are “wetted down” as they enter the cave to limit the organic matter shed and left behind


Painted Fiberglass study

As explained above, soon after installation, cave workers noticed the development of slime biofilm on painted fiberglass surfaces used to hide electrical and plumbing equipment.   Our hypothesis was that the paint on the fiberglass serves as a source of carbon and energy for naturally-occurring cave isolates which grow copiously in comparison to the rest of the community.  Further, we hypothesized that these isolates produce slime to (i) aid in biofilm formation, and (ii) protect against paint toxicity.

Figure 3. (above) A comparison of growth taken from a swab taken from a painted fiberglass surface containing a slimy biofilm (left) and from the floor of the cave.

Figure 3.

 

To test the hypothesis, the nine isolates cultured from the painted fiberglass were grown on a mineral salts medium agar containing acrylic paint as the sole carbon and energy source (see Table 1 below).  Of these nine isolates, several grew on the paint-agar plates even at high paint concentrations.  Three isolates that could grow on the paint-agar plates also produced slime during growth.  Two of these isolates have closest relatives that are known slime producers:

Table 1. The ability of isolates from painted fiberglass surfaces to grow on acrylic paint as the sole source of carbon and energy.

Isolate ID

Mucoid Colony

Growth on paint
(mg/L)

100

1,000

10,000

PF-B
PF-H
PF-I
PF-A
PF-G
PF-M
PF-D
PF-K
PF-F

Yes
Yes
Yes
No
No
No
No
No
No

+
+
+
+
-
+
+
+
-

+
+
+
+
-
-
+
-
-

+
+
+
-
-
+
-
-
-

Research outcome:  Instead of fiberglass, concrete was used in the development of the Big Room (the second room in Kartchner that was developed for tourism)

updated 7/2009

 

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