The potential hazards associated with the genes introduced in the GM papayas transferring to animals, including humans, could be highly variable, broadly depending upon the phenotype of the recipient and any changes to the survival or reproductive capacity of it or its progeny. The potential hazards posed by the specific gene sequences, are as follows:
Animals could become resistant to the antibiotics. If the transfer occurred to humans or other animals treated with these antibiotics, the antibiotics may be inactivated before they are able to control the targeted bacterial pathogen. The possibility of the transfer of these genes reducing the efficacy of antibiotic treatments has been considered.
GUS reporter gene:
This would not present a risk to human health or the environment, in the extremely unlikely event that it occurred.
CaMV 35S promoter and other regulatory sequences:
The expression of endogenous genes in the recipient animal could be altered. If a change in normal gene expression did occur, the hazard to the recipient animal and to the environment would depend on the specifics of the resultant phenotypic change.
Section 3.2 Likelihood of gene transfer from the GM papayas to animals
The most significant route for entry of foreign DNA into humans is through food, as it passes through the gastrointestinal tract. The epithelial lining of the gastrointestinal tract is routinely exposed to foreign DNA released from food. The likelihood of DNA from the GM papayas proposed for release transfering to humans directly, following consumption is extremely unlikely.
In addition, it should be noted that Food Standards Australia New Zealand (FSANZ), is responsible for human food safety assessment. Currently, the applicant has not applied to FSANZ for evaluation of material from the GM papayas for use in human food. FSANZ approval would need to be obtained before it could be used in human food.
Transfer to humans via bacteria
It has been hypothesised that the genes introduced to GM plants could be transferred to humans (and other animals) indirectly, via bacteria that occur in the gut. Several studies have investigated the likelihood of this potential hazard, since the gastrointestinal tract may be exposed to DNA from ingested food for many hours on a daily basis and microorganisms colonise the length of the gastrointestinal tract, aiding the digestive process.
Netherwood et al. (2002) investigated whether DNA in food was able to survive the human gastrointestinal tract. Seven ileostomists (people with a colostomy bag) were recruited for the trial in which they were fed GM (Roundup Ready) soya products. The survival of both the Roundup Ready gene and an endogenous soya gene were traced. Surprisingly, a large portion of the transgene from the GM soya was observed to survive the passage of the small intestine. The level of persistence of the endogenous soya gene was similar to that of the Roundup Ready gene, indicating that the transgene was degraded at a similar rate to the bulk soya DNA. In this study, GM soya was used as a model experimental system and the results have broad applicability to many other GM plant foods, including GM papaya.
To determine whether the transgene in the GM soya survived passage through the complete gastrointestinal tract, a further 12 human volunteers were fed the GM soya-containing meal. The transgene was not detectable in faeces from any of the subjects. These data indicate that GM soya, although surviving passage through the small intestine, is completely digested in the large intestine and the colon. It is possible that the small intestine of ileostomists differ from that in people with an intact gastrointestinal tract. For example ileostomists could secrete lower levels of DNAase. Alternatively, the rate of passage of the digesta could differ, or the structure of the microbial ecosystem in the small intestine may be quite different.
Microorganisms present in the digesta samples from the ileostomists were tested for evidence of gene transfer from the GM soya. Genetic tests for the presence of the DNA to bacteria confirmed that gene transfer had occurred at a very low levels. However, individual bacteria harbouring the transferred DNA could not be isolated, indicating that the bacteria containing the GM soya transgene represented an extremely minor component of the intestinal microflora of these subjects.
Studies on intestinal colonising bacteria such as lactabacilli, and Salmonella typhimurium (an intracellular pathogen) indicate that transfer of plant genes in food, specifically transgenes in soya foods, to the intestinal epithelium, is unlikely to occur because gene transfer could not be induced under highly selective experimental conditions.
Potentially, tissues from the GM papayas proposed for release, including the fruit, may be fed to farm animals, exposing their gastrointestinal tract to the introduced genes. The fate of DNA in the digestive tract of various animals has been studied. A review of the safety issues associated with the DNA in animal feed derived from GM crops (Beever & Kemp 2000) indicated exposure to introduced DNA from GM crop material is negligible compared with normal exposure to non-GM DNA. They calculated that in a diet containing 40% GM maize, the introduced genes would represent 0.00042% of total dietary DNA intake.
Using GM glyphosate-tolerant canola as a model experimental system, Alexander et al. (2002) investigated the digestive fate of DNA from GM plants. They used PCR to detect the presence of two genes in various canola feed fractions following in vitro incubated in bovine ruminal fluid. The genes analysed were the CP4 EPSPS gene introduced by genetic modification and an endogenous nuclear-encoded rbcS gene (encoding the small subunit of the photosynthetic enzyme Rubisco).
Whole seed, cracked seed, canola meal or a prepared diet (containing 6.5% canola meal) were examined. Processing of canola seed to meal was found to significantly reduce the amount of DNA detected. There were no significant differences in the detection of the introduced or endogenous gene. These feeds were incubated in batch cultures of ruminal fluid. Both genes could be detected in the cultures of whole and cracked seed for up to 48 hours, but only up to eight hours for meal and four hours for the prepared diet. The genes were detected in the plant debris but not in the aqueous phase of the ruminal cultures. The authors concluded that the plant DNA was rapidly degraded by rumen fluid and that the persistence of DNA was inversely related to plant cell digestion (Alexander et al. 2002). These results support the conclusion that the rapid degradation of DNA following release from plant cells during ruminant digestion represents a considerable barrier to transfer of plant DNA, GM and non-GM, to rumen bacteria or to ruminant animals.
Einspanier et al. (2001) investigated the fate of DNA from GM maize fed to cattle and chickens, using PCR to detect the introduced cryIA(b) gene (which confers resistance to insects) and an endogenous plant chloroplast gene. Since multiple chloroplasts are present in plant cells, more copies of the chloroplast gene are in the GM maize than of the cryIA(b) gene.
For cattle fed GM maize silage, both the cryIA(b) gene and the chloroplast marker were detected in chyme (duodenal juice). The chloroplast marker was detected in lymphocytes and faint signals were occasionally detected in milk, but it was not detected in faeces, whole blood, muscle, liver or spleen. The cryIA(b) gene was not detected in any of these samples (Einspanier et al. 2001).
In chickens fed a diet containing GM maize, the chloroplast marker was detected in muscle, liver, spleen and kidney, but not in faeces or eggs. In contrast, the cryIA(b) gene was not detected in any tissue sample or eggs (Einspanier et al. 2001).
The possibility of DNA transfer in the gut has also been investigated by feeding mice large quantities of purified bacteriophage DNA (Schubbert et al. 1997). Bacteriophage DNA was detected in the faeces and the livers of mice as well as in rarely in newborn mice (Schubbert et al. 1997). However the relevance of this work to gene transfer from GM plants was questioned by Beever and Kemp (2000), who concluded that the bacteriophage DNA was in a form which would stimulate a response by cells of the immune system, and that the cells containing this DNA in various organs and newborns were macrophages involved in scavenging and removing foreign DNA.
In the rare event of plant DNA uptake by animals cells, a further step of chromosomal integration has not been demonstrated. Furthermore, any uptake of plant DNA is likely to occur in non-reproductive (somatic) cells such as immune system or gut epithelium cells, and the introduced gene would not be transmitted to progeny.