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Ten years of Bt maize cultivation: Horizontal gene transfer of no significance

Scientists from France and Switzerland have been studying soil bacteria from a field where genetically modified Bt maize has been growing for 10 years. They wanted to find out whether controversial antibiotic-resistance genes can in fact transfer from transgenic plants to bacteria, as is widely feared. They have concluded that transgenic plants play no part in the spread of antibiotic resistances.

Bacteria have special mechanisms which enable them to exchange genetic information directly without sexual reproduction. For this reason it is feared that antibiotic-resistance genes that are used as marker genes in transgenic plants could be absorbed by pathogenic bacteria and so reduce the effectiveness of important antibiotic drugs. Antibiotics are widely used in human and veterinary medicine and for a long time they were also added to animal food to promote animal growth and performance. This has led to the emergence of bacterial resistances to antibiotics used in medicine. The question is, do genetically modified plants also help spread this kind of antibiotic resistance?

“The risk should be regarded as almost zero.” Pascal Simonet of the Ecole Centrale de Lyon and his colleagues have investigated whether transgenic plants contribute to the spread of antibiotic resistances.

Bt176 maize in the trial field at Bazi├Ęges near Toulouse

Fig.1 Resistance pattern of ampicillin-resistant bacteria (a-g). None of the bacteria were resistant only to ampicillin and carbenicillin, unlike genetically modified E. coli bacteria with the Bt176 plasmid

Leaf pieces infected with Agrobacterium are grown on a selection medium. This culture medium contains an antibiotic on which only those cells in which the transformation has been successful can survive.

Antibiotic-resistance genes are often inserted into plants in addition to the actual target gene. Following genetic transformation, the plant material (apples in this example) is placed on an antibiotic-rich culture medium. Only those plant cells which have been successfully transformed will survive and grow on this medium, since they are resistant to the antibiotic.

Proportion of ampicillin resistant bacteria *

Bt176 soil Conven tional maize soil Prairie soil
0.4- 6.5 % 5.5- 8.0 % 54.4- 69.6 %

*as a proportion of the number of cultivable bacteria

Gene transfer from plant DNA to bacteria is considered to be highly unlikely because a whole series of conditions are required before it can occur at all. As yet, this type of horizontal gene transfer has not been detected under field conditions. Even in the laboratory, it could only be provoked with the help of specially constructed recipient bacteria.

To assess the likelihood and the significance of a possible transfer of antibiotic- resistance genes from transgenic plants to bacteria, scientists from France and Switzerland have studied soil bacteria from a field in south western France where genetically modified Bt176 maize has been growing for 10 years. By way of comparison, soil samples from a conventional maize field and from uncultivated land (prairie soil) were also investigated.

Bt176 maize contains a “bla gene” (blaTEM116) as a marker gene, in addition to the gene which makes it resistant to the corn borer. This gene confers ampicillin and carbenicillin resistance by producing a specific beta-lactamase enzyme. It is one of the most commonly occurring bla genes from a whole family of beta-lactamases. The corresponding antibiotics, which include penicillin, are the largest group of antibiotics used in medicine.

Widespread ampicillin resistance

The researchers initially studied only those bacteria from the soil samples which could be propagated on a culture medium. These account for less than 1 percent of soil micro-organisms. Between 0.4 and 8 percent of these bacteria from the soil samples collected from agricultural land were found to be resistant to ampicillin. It made no difference whether conventional or Bt maize had been grown on the fields. However, the number of resistant bacteria varied significantly between the cultivated soils on the one hand and the prairie soil on the other, which had a very high proportion of resistant bacteria. According to the scientists, this is an indication that bacterial communities which are not affected by farming practices have a higher proportion of naturally occurring antibiotic resistances. Bacteria which produce antibiotics themselves often carry resistance genes for their own protection.

To discover the origin of the resistance genes, the resistant bacteria identified were then exposed to various antibiotics and their resistance patterns were recorded. Most of the 576 bacteria isolated were resistant to several antibiotics (see Fig.1)

The resistance genes identified were then examined in more detail using molecular-biological methods (PCR). A bla gene was identified in 505 of the 576 bacteria (87.7 percent). According to the authors, this indicates the natural preference for these genes amongst ampicillin-resistant soil bacteria.

Eighty of the PCR results were broken down further. Among other things, ten blaTEM116 genes were found, distributed over all the soil types. This indicates that the ampicillin-resistance gene, which was used for Bt176, is also found in soils where no transgenic plants have been grown. This is not surprising, because the gene for the genetic transformation was isolated from soil bacteria. The fact that blaTEM genes were also found in the prairie soil 400 kilometres away confirms the prevalence of these genes.

In addition, the large number of non-cultivable bacteria was also studied using molecular-biological methods. More than 150 different blaTEM genes were identified.

Bacterial diversity

The diversity and composition of the bacterial communities in the different soil samples were also investigated to find out whether the occurrence of similar bla genes implies similar microbial communities. This is apparently not the case. There were significant differences in the composition of the micro-organism communities in the three soils, with the greatest difference occurring between the maize fields and the uncultivated land. Possible changes resulting from the cultivation of Bt maize are therefore less significant than the changes resulting from soil type, plant growth stage, variety or crop differences.

Horizontal gene transfer - no risk

Even if horizontal gene transfer is possible in principle, the authors believe that it is of no significance to microbial communities in the soil. They claim that the cultivation of transgenic plants for more than 10 years in one field has had no measurable effect on the occurrence of antibiotic-resistances and their spectrum. They believe that this is largely due to the fact that the genes are already commonly found in the soil. Horizontal gene transfer from transgenic plant DNA to bacteria is so rare that it could not contribute to a further increase in the widespread antibiotic resistance which already occurs naturally in bacteria.