Some Japanese scientists believe that the Wagatsuma agar plate, containing human red blood cells, can determine the virulence of a strain of Vibrio parahaemolyticus. This hemolytic test, referred to as the Kanagawa phenomenon, is based on the ability of an organism to digest blood cells within 24 hours. The Wagatsuma plate (top left photo) illustrates this. Cleared zones around the two colonies at top show digested blood cells, indicating pathogenicity, and the colony at bottom, without a cleared zone, indicates

nonpathogenicity. In a serological test (bottom)

left photo) to confirm a particular strain of Vibrio parahaemolyticus, a saline solution containing the organism is mixed with antiserum specific for it. The test is postive if agglutination takes place, as shown at right, and negative if no agglutination takes place, as shown at left. At top right is shown a pure culture of Vibrio parahaemolyticus streaked for colony isolation on a selective agar plate. This electron photomicrograph (bottom right) of Vibrio parahaemolyticus, made by Dr. Rita Colwell, Georgetown University, shows the organism at a magnification of around 108,000.

about similar outbreaks in the United States. On the face of it, the problem appeared to be confined to Japan. But by 1968, Drs. John Liston and John Baross of the University of Washington reported the isolation of Vibrio parahaemolyticus from fish, shellfish, and crustaceans and from the waters and sediments-of the Puget Sound area of the Pacific Coast. Thus the hazardous potential had assumed more serious proportions.

The Division of Microbiology bacteriologists and the other scientists of the Food and Drug Administration provide the scientific support that makes it possible for this Agency to assure the safety of the food supply under the Food, Drug, and Cosmetic Act. It is the Division's task to be alert to new potential hazards such as toxic substances produced by micro-organisms or to infectious bacteria capable of causing foodborne disease, and to initiate studies to determine their possible significance. The recovery of Vibrio parahaemolyticus from the Puget Sound area introduced a new health problem, with several unknowns.

Many questions arose simultaneously. Was the organism confined to the Puget Sound area or could it be found on the rest of the Pacific Coast? Was it present on the Gulf Coast and the Atlantic Seaboard? What marine forms may be involved as hosts? What were the quantitative relationships between this micro-organism and the disease it caused? Had there been outbreaks of illness from eating seafood in the past that had been attributed to other causes? What about the pathogenicity of the Puget Sound isolates? Japanese research had shown that there were both virulent and avirulent forms of this organism. It would be necessary for FDA to get the answers to many of these questions as soon as possible, and Division microbiologists set about the task.

Japanese scientists found that the virulent forms were recovered invariably from patients ill with the disease and the avirulent forms mainly from marine animals, water, and sediment samples instead of human victims. They also observed a positive correlation between enteropathogenicity and the Kanagawa phenomenon. The Kanagawa reaction is based on the capability of Vibrio parahaemolyticus to produce a specific type of hemolysin (beta type) when incubated 24 hours on a specially prepared blood agar plate. At the present time no one can explain satisfactorily whether the avirulent types of Vibrio parahaemolyticus found in seafood may be converted into the infectious types capable of producing disease outbreaks in man and, if such a conversion takes place, what would cause it.

A considerable amount of work remains to be done by scientists to delineate the pathogenic role of this organism. Dr. Sakazaki has reported that a highly purified Kanagawa hemolysin (produced by Dr. Y. Obara) failed to elicit a pathogenic response in the ligated ileal loop portion of the intestine of test rabbits. This puri

fied substance, however, was lethally toxic when inoculated into the peritoneal cavity of test mice. In FDA's Division of Microbiology laboratories, different fractions of crude Kanagawa hemolysin have proved lethal to mice when inoculated intraperitoneally. A pure enterotoxin substance, if one indeed exists, has yet to be isolated, purified, and used to produce the specific disease syndromes in either man or animals.

As part of a program to intensify and expand awareness and knowledge among public health officials in this country about Vibrio parahaemolyticus as a pathogen with the same potential for causing disease here as in Japan, the Division of Microbiology arranged for a month's visit by Dr. Sakazaki (see photos) to the United States in July-August 1971, under the sponsorship of the World Health Organization, for a series of seminars to bring Federal, State, and university microbiologists up to date on the newest findings in Japan concerning this organism.

Acting as a temporary consultant to the Division, Dr. Sakazaki presented seminars on Japanese research to audiences in Washington, Cincinnati, Seattle, and College Station, Texas. Ironically, Dr. Sakazaki's visit to this country ended just a few days before the Maryland outbreak became the first confirmed instance of illness caused by Vibrio parahaemolyticus in the United States.

In the United States, one unusual pathogenic aspect of Vibrio parahaemolyticus has been observed that had not been reported in the scientific literature before. For some time now, Dr. R. E. Weaver of the Center for Disease Control at Atlanta has been collecting a number of cultures of the micro-organism that have been isolated from infections of the hands and feet, eyes and ears, and also from the bloodstreams of persons who have been in contact with marine shore areas of different parts of the United States. These infections have usually started from scratches or cuts caused by the sharp edges of clams or oyster shells embedded in the sand.

The serology of Vibrio parahaemolyticus has been well worked out by Japanese scientists. The microorganism possesses three antigenic components-the O, K, and H antigens. The O antigens are considered to be body or somatic in origin and are stable in the presence of heat. The K antigens are derived from the capsular or envelope portion of the bacterium and are primarily polysaccharide in nature. The H or flagellar antigens are of little value in serological identification since these are nonspecific in the bacteria genus Vibrio.

By the most recent count, there are 11 O group antigens and 53 K antigens associated with this microorganism. This results in the current recognition of 54 individual serotypes of Vibrio parahaemolyticus-based on the combination of O and K antigens found to occur naturally in this bacterium. When the Japanese investigators first devised this antigenic scheme, the

micro-organisms used were obtained from patients only. But since there are still many undescribed serotypes of Vibrio parahaemolyticus in seafoods that evidently continue to infect humans, the number of newly found serotypes continues to increase. In our own experience at the Division of Microbiology laboratories with hundreds of seafood isolates, we have been able to serotype only 40 percent, the other 60 percent being untypable at the present stage of technical capability.

Scientists in the United States, since the findings by Drs. Liston and Baross reported in 1968, have been active in isolating and identifying the micro-organism from the marine environment and studying its properties. In FDA's Seattle District, Dr. Gary Houghtby is studying the response or growth of the micro-organism at different temperatures. In the San Francisco District, Warren Landry has been isolating the bacterium from the coastal shellfish of Texas and Louisiana for the past two years. Mr. Landry also has demonstrated that it may be recovered from frozen shellfish products for up to three months after freezing. Drs. Carl Vanderzant and R. Nickelson of the Texas A & M University have recovered the bacterium from commercial shrimp pond culture operations and also are making methodology and serology studies.

On the eastern seaboard, Dr. Morris Fishbein and Barry Wentz, both in FDA s Division of Microbiology, have developed the methodology of isolation and recovered hundreds of strains of the micro-organism from many varieties of marine fish, shellfish, and crustaceans in Chesapeake Bay and all the coastal areas of the United States, and are now working on the Kanagawa hemolysin. In the Division's laboratories at Cincinnati,

Dr. Robert Twedt and his associates have studied the rabbit ileal loop response to this organism. More recently they have devised a culture medium for use in recovery of Vibrio parahaemolyticus.

Another pioneer in this country is Dr. Rita Colwell of Georgetown University in Washington. She and her microbiology students have conducted an extensive study of the nearby Chesapeake Bay area and have recovered many Vibrio parahaemolyticus strains. Her papers on the taxonomy of the micro-organism are a valued contribution in the field. In FDA's Boston District, Barbara McEachern has recovered the bacterium from a variety of seafoods taken from the New England shore area. Likewise, Dr. W. K. Thomson of the Canadian Food and Drug Directorate has made a number of isolations from seafood and currently is making pathogenicity studies.

Because of the still limited knowledge about the prevalence of Vibrio parahaemolyticus foodborne infection in the United States, there is a need to collect enough information to assess its relative importance as a foodborne disease here. FDA's Divisions of Microbiology and Shellfish Sanitation and other scientists in the Federal and State agencies and universities in this country, Japan, and Canada are working in close conjunction to keep abreast of the continuing developments concerning Vibrio parahaemolyticus and the disease it causes. In this regard, all State and municipal health department laboratories should include among their various tests of food suspected of causing an outbreak an examination for the presence of V. parahaemolyticus. Appropriate procedures for this purpose are available from FDA's Division of Microbiology.

ATLANTA DISTRICT Professor James Harvey Young, Department of History, Emory University, Atlanta, author of the books The Toadstool Millionaires and The Medical Messiahs, and Professor Chikataro Kawasaki, once associated with the University of Osaka School of Pharmaceutical Sciences and now in the process of establishing a school of pharmaceutical sciences at Kobe Gakuin University, visited Atlanta District on June 23. They discussed the operation of FDA with Acting Regional Food and Drug Director Joseph J. Milunas and Laboratory Director Sol Cohen and toured the office and laboratory facilities.

Data processing was one of the major topics discussed at the Association of Food and Drug Officials of Southern States meeting in Lexington, Kentucky, on April 18. Atlanta District personnel explained FDA's current system of data retrieval and the proposed system to become effective in July 1972. State food and drug officials seemed to agree that standard industrial classification would appear to be the most compatible identification system. General meetings as well as formal and informal conferences with State personnel indicated that FDA officials are making progress in their attempt to work more closely with their State counterparts.

BOSTON DISTRICT A training session in State shellfish program evaluation was held June 8-11 at FDA's Northeast Technical Services Unit (NTSU) in Davisville, Rhode Island. All regional shellfish consultants, special program branch chiefs, and various headquarters personnel attended the training program, conducted by the Division of Shellfish Sanitation, Office of Food Sanitation, Bureau of Foods, in cooperation with FDA Training Institute and NTSU.

The course is designed specifically for regional shellfish consultants and other FDA personnel responsible for the review and evaluation of State shellfish control programs. Its purposes are to provide detailed instructions in policy and procedures for uniform evaluation of programs, to evaluate problems associated with specific geographical localities, and to present the relationships of the shellfish division with other Federal agencies. Boston District was visited on June 30 by eight members of a Japanese delegation interested in the analysis of canned tunafish. Analyst Tony LaTerza demonstrated the procedure for examining canned tuna for decomposition and its preparation for mercury analysis. The procedure for identifying mercury in fish was also demonstrated. Frank Allhands, Office of Product Technology, Bureau of Foods, Washington, and several top District personnel also participated in the six-hour meeting.

CINCINNATI DISTRICT A seizure was made of 310,000 units of fireworks, valued at $35,000, by U.S. marshals and agents of the Internal Revenue Service on June 25 at Miller Fireworks and Novelty Co., Inc., Holland, Ohio. The firm was the source of a majority of fireworks found at 55 retail outlets which had been charged with selling illegal fireworks. The firm's owner, John F. Miller, was arrested by Internal Revenue Service agents and charged with illegal purchase of black powder and illegal manufacture of explosives. Cincinnati District assisted in the action.

Cincinnati Consumer Specialist Catherine Knarr, as a member of the Greater Cincinnati Nutrition Committee, is helping in a survey of various health departments and other agencies with nutritional responsibilities. The survey's purpose is to assess current services available in the greater Cincinnati area and the nutritional problems and needs of the area so that resources, activities, and services of the various agencies can be coordinated. Cincinnati District is continuing a series of meetings on equal employment opportunities for District personel. Speakers have included Voyce Whitley, FDA, EEO director; Dewey Fuller, assistant director, Cincinnati Urban League; and Dr. Edith Parkey, past president, Federally Employed Women, and former consultant to the Post Office Department as architectural engineer.

KANSAS CITY DISTRICT Accidental spilling of parathion, an extremely toxic pesticide chemical, inside a commercial truck at Omaha, Nebraska, on June 29 resulted in the destruction by burial of approximately $10,000 worth of candy, paper goods, school supplies, and bedding contaminated by the chemical. The truck, enroute to Omaha, picked up a cargo of some 70 cases of candy at a St. Joseph, Missouri, candy manufacturer. The trucker then added to the load various chemicals, including parathion, from a St. Joseph chemical company. When the truck was opened at Omaha, it was found that a 30-gallon drum of parathion had ruptured and contaminated the cargo. Four employees were exposed to the parathion and were placed under observation. The trucking firm assumed full responsibility for the incident and offered its cooperation in the investigation.

Involved in the investigation and supervision of destruction of the contaminated items were FDA's Omaha resident inspector; U.S. Department of Transportation personnel; the Nebraska Department of Agriculture, Dairy and Food Division; and an industryformed Pesticide Safety Team headed by the Stauffer Chemical Co., Omaha.