From the 1975 Masters Thesis: Bacteriological and ecological observations on the northern pitcher plant, Sarracenia purpurea. John A. Lindquist. Department of Bacteriology, University of Wisconsin, Madison, WI.
In the study of plant-microbial relationships outside of plant diseases, most research has been done on the rhizosphere, especially concerning nitrogen fixation. The leaf ecosystem has been studied relatively little, but interest and research on the "phyllosphere" has increased in recent years. An international symposium on epiphytic microorganisms has recently resulted in an integrated and wide-ranging collection of papers (Preece and Dickinson, 1971); nowhere, however, are carnivorous plants and their bacteria discussed.
There are various factors influencing the amount and diversity of microorganisms on leaf surfaces including moisture, temperature, leachates, condition of the leaf and airborne micro-organisms (Sinha, 1971). Moisture is necessary for extensive colonization of the leaf surface, especially on tropical plants which grow in humid, high-rainfall environments (Brock, 1966). White (1943), working on relatively dry bean vines found 100% of excised vine tips developed mold when placed in nutrient, but only about 50% developed bacteria.
There may be many microbial interactions on plant surfaces, some antagonistic (e.g. fungi being depressed by bacteria) and some mutually favorable (Klincare, Kreslina and Mishke, 1971; Fraser, 1971). Epiphytic bacteria may indirectly benefit the plant, such as nitrogen-fixing bacteria which may pass nutrients to the plant, especially upon their death. Ruinen (1956) and Centifanto and Silver (1964) respectively discovered Beijerinckia and Klebsiella growing and fixing nitrogen on the leaves of tropical plants; Beijerinckia had been previously found only in soil.
Of the bacteria growing on leaves, a high proportion are chromogenic; perhaps the best known of these is Erwinia herbicola, now designated as Enterobacter agglomerans by clinical microbiologists (Crosse, 1971; Ewing, 1972). Bacterial genera found on a wide variety of plants have included the following: "Aerobacter," Corynebacterium, Flavobacterium, Lactobacillus, Pseudomonas, Chromobacterium, Micrococcus, "Sarcina", Mycobacterium and Bacillus. Actinomycetes and yeasts are also frequently found (Crosse, 1971; Klincare, Kreslina and Mishke, 1971).
Only two papers in this century have contained original experimental work on the bacteria found in the North American pitcher plants.
Hepburn and St. John (1927) studied fifty closed pitchers of various species of Sarracenia (including S. purpurea) and Darlingtonia and found no evidence of bacteria in the pitcher cavities. The cavity contents were inoculated onto "plain nutrient agar" and incubated at 37°C. One could argue that a lower, more realistic temperature could have been used as well as a diversity of media to produce a valid conclusion on sterility.
Hepburn and St. John (1927) also studied 39 open pitchers of these species, each pitcher containing prey. The authors transferred the contents of the pitchers to plain nutrient agar slants and then inoculated the grown, mixed slant cultures to media to test for proteolytic, alkali-forming, acid-forming and "colon-aerogenes" bacteria; all media were incubated at 37°C. Proteolytic bacteria were generally found, and an alkaline reaction was usually produced from the nitrogenous substrates used to detect alkali-formation: ammonium lactate, ammonium tartrate, acetamide, urea, asparagine and glycocoll. In the media for detection of acid production from lactose and glucose, the cultures showed a marked tendency to attack the peptone component of each medium, forming alkaline rather than acid reactions. Most of the pitchers (none for D. californica) were found to contain "colon-aerogenes" bacteria, and it was concluded that these bacteria were contributed by insect prey. However, the modern distinction of fecal coliforms (those gram-negative rods which actively produce gas from lactose fermentation at 44.5-45.5°C) vs. total coliforms (gas at 35-37°C) was not made, and thus it is not possible to judge whether the authors dealt with the true intestinal coliform, Escherichia coli, or with the coliforms which occur naturally on plants (Enterobacter, Klebsiella).
Because the bacteria digested the protein "so slowly" in these experiments (at 37°C), it was stated that "their part in the digestion of the prey must be a minor one in the genus Sarracenia, the protease of the pitcher liquor [probably meaning a plant-produced protease] playing the leading role." Tissue enzymes of the captured insects were also believed involved. The strong bias of Hepburn and St. John (1927) in favor of plant protease and against bacterial protease persisted in the subsequent literature concerning digestion in pitcher plants.
Plummer and Jackson (1963) studied the bacteria of S. flava with plate counts on nutrient agar, gram stains and pH determinations (discussed previously) of the pitcher fluid during digestion of meat and insects, both in vitro and in vivo. An increase in the total number of bacteria with a decrease in the variety of dominant colonial types was observed from the plate counts during the course of digestion, coincident with an increase in the relative number of gram-negative bacteria observed on the slides. Maximum abundance of bacteria occurred between 6 and 12 days after the initiation of digestion. Rods (including spore-formers), cocci and yeasts were observed throughout the course of digestion; no filamentous fungi or streptococci were seen. No isolations or identifications were made, however, gram-negative bacteria similar to those often associated with the putrefaction of meat and organic waste (Pseudomonas, Achromobacter, Proteus and gram-variable Clostridium spp.) were suspected.
Hydrolysis by acid and plant-produced proteolytic enzymes were thought by Plummer and Jackson (1963) to precede the hydrolytic action of bacteria which later, through proteolysis and subsequent ammonification, create alkaline conditions. As opposed to Hepburn and St. John (1927), Plummer and Jackson (1963) considered bacteria to be significant and active in the digestive process, especially with materials having a relatively large surface area. As macerated insects were used in this study, the roles of bacteria and insect autolytic enzymes were considered more immediately significant under the experimental conditions than would be the case in the field.
It should be appreciated that the S. purpurea pitcher fluid is probably very conducive to the growth and activity of proteolytic bacteria; hence, a digestive secretion may not be necessary. However, for S. flava pitchers, into which rain enters with difficulty, the importance of a plant-produced digestive secretion has been indicated.
Go back to Part I of the Literature Review.
Go back to Part II of the Literature Review.
Go to the References.
Return to the Pitcher Plant Project "Index" Page.
Page last modified on 1/9/98 at 9:45 AM, CST.
John Lindquist, Department of Bacteriology, University of Wisconsin – Madison