By David N. Leff

See if these statistics from the World Health Organization surprise you:

In 1996, the number of people who died the world over was 52 million. Of that total, the No. 1 global killer was microbial infections, which took 17 million lives.

¿Close behind,¿ observed molecular geneticist Michael Mahan, ¿came circulatory diseases ¿ heart attacks and strokes ¿ with 15 million deaths. This was followed by cancer, with 6 million. So, infectious diseases outnumbered cancer almost three to one.¿

Mahan is an associate professor of genetics, focused on pathogenesis, at the University of California in Santa Barbara.

¿Worldwide,¿ he said, ¿but not in the U.S., deaths from typhoid fever caused by the Salmonella typhi bacterium is a problem. However, 4 million Americans a year get Salmonella food poisoning from a somewhat less virulent strain, Salmonella typhimurium. That leads to billions of dollars in medical care, lost wages and products.¿ (See BioWorld Today, Dec. 29, 1998, p. 1.)

Much of that food poisoning comes from bacterially contaminated beef and poultry. ¿Salmonella,¿ Mahan said, ¿is as much an animal pathogen as a human one. And it¿s a big problem in the food industry. When you go down to the store and buy your chicken, many of the birds are contaminated. Which is why you have to cook your chicken very thoroughly, or run the risk of getting food poisoning.¿

Mahan has just announced discovery of a putative oral vaccine against Salmonella typhimurium which, he told Bio World Today, ¿we see as having an immediate impact on the poultry industry. Our idea is to spray the vaccine on the chicken feed and let the chicks eat it.¿ His research report in the current issue of Science, dated May 7, 1999, is titled: ¿An essential role for DNA adenine methylation in bacterial virulence.¿ A poultry vaccine is the least of what the article covers.

DNA adenine methylation describes a gene better known as Dam. ¿Its protein, DAM, has been known for a long time, but so far only in Salmonella bacteria,¿ Mahan said. ¿What¿s new about our paper is it¿s the first report that Dam has anything to do with pathogenesis. And what accounts for some of the excitement is that this known Salmonella gene might also exist in many other infective pathogens that cause serious health problems. These include, amongst many others, the deadly organisms of cholera, dysentery, meningitis, typhoid fever, the plague.¿

Salmonella itself numbers upwards of 2,500 strains, so the team¿s first effort is to determine if Dam, and the myriad slave genes it commands in the bacterial cell, are limited to the one strain they have explored. ¿What we don¿t know,¿ Mahan said, ¿is whether this gene acts as a master switch, controlling bacterial infections in other organisms, as it presumably does in Salmonella. If it does, we believe this will lead to a new generation of vaccines and antibiotics. We are working on that right now, furiously.¿

The first thing Mahan and his co-authors did was create a Dam-minus Salmonella knockout bacterium, thus deprived of its DAM enzyme. ¿When we genetically knocked out the DNA encoding that enzyme,¿ Mahan said, ¿we had a mutant of the Dam gene that encodes DAM. What happened was that in the laboratory it lost the expression of many of its bacterial weapons. So, this enzyme seems to be clued in to a specific class of genes that we have discovered. The two main points of our findings are that we¿ve identified a master-switch controlling bacterial infection, and utilized this information to make a vaccine which has proved to be protective in mice against Salmonella infection.¿

Briefly, when the team vaccinated 25 mice with its mutant Salmonella vaccine, then challenged them with 10,000 times the lethal dose of wild-type bacteria, all 25 animals survived with nary a symptom of infection. But of 12 control animals denied immunization, all 12 died swiftly.

Salmonella are infamous for evading the mammalian immune system. Their camouflage consists, Mahan suggested, of expressing their virulence genes only at the last moment before infection, thus denying the immune defenses a shot at their targets until too late.

¿The bottom line,¿ he suggested, ¿is that the bacteria hide from the immune system. And when Dam is knocked out, they reveal ¿ since they¿re global gene-regulating villains ¿ many of the gene products. These include specific adhesion sequences that allow them to stick to host cells, and toxins that allow them to destroy host cells.

¿In wild-type organisms, these are expressed only at a specific time, at a specific place during infection,¿ Mahan said. ¿You can¿t fight what you can¿t see. Now, the mutants are expressed and showing their hand all of the time. Bacteria are great poker players, but no matter how good at poker you are, if you lay your cards down face up, you¿re dead. That¿s why we get such a great immune response, because the mutant organism is showing everything to the immune system.¿

New Company Seeks Name, Financing, Partner

Mahan expects it will take two years to get his poultry vaccine to market. ¿There are certain modifications for safety that we have to make, in addition to having a single mutation that cripples the bacterium and also induces immunity,¿ he said. ¿What we don¿t want is having this mutated master switch revert to wild type. If it did, we wouldn¿t be immunizing chickens, we¿d be infecting them. So, we¿re proceeding with those modifications as feverishly as we can.¿

An eventual human-protecting vaccine awaits further basic research.

¿We co-authors are founding a company to bring the technology into practice,¿ Mahan said. ¿It would make vaccines against many of these diarrheal pathogens. That¿s going to be the initial force. And the second time line is to make antimicrobials. At some point, we¿ll probably have to partner up with some big company to do that, because you have to screen hundreds of thousands of compounds against this enzyme. Right now, we¿re doing the corporate searches, dotting the i¿s and [crossing the] t¿s on our future firm¿s name, and scrambling to get it going.¿

The patents are owned by the university, he added, which ¿never relinquishes its rights to intellectual property, so they¿re in the driver¿s seat.¿

As for financing, ¿we haven¿t nailed that down,¿ Mahan concluded. ¿We felt that when this paper came out, we would have options that we didn¿t have last week. And our phones are ringing off the hook today, including some calls from the pharmaceutical area. So, we¿re seeing spurts of excitement in the applied¿ sector ¿ to put it diplomatically.¿ n