Microbiology Mary Jane Ferraro and Robert C

Chapter  Microbiology Mary Jane Ferraro and Robert C. Moellering Jr.

he history of Microbiology (or Bacte- abrupt departure the following year, Dr. Law- Triology, as it was originally known) at the rence Kunz, who had come to the MGH as an Massachusetts General Hospital (MGH) is a Assistant Bacteriologist in 1952, assumed de facto complex one. Although Dr. Reginald Heber responsibility for the laboratory until he was offi - Fitz’s pioneering work established the pathogen- cially appointed chief in 1956. During his tenure esis of appendicitis in 1886 (15, 41; and see chapter the Department of Bacteriology developed close 2), the laboratory specialty of microbiology owes ties with the Infectious Diseases Division and the its origins to Dr. James Homer Wright, who was Department of Medicine. Th is was also a time appointed Pathologist at MGH in 1896. Wright during which a number of “boutique” clinical was involved in a number of landmark studies laboratories sprang up throughout the hospital. in the etiology of parasitic and other infections, Microbiology and Infectious Diseases were no established the fi rst diagnostic bacteriology labo- exception, and in rapid order independent laboratory functions at the MGH, and hired several ratories of Antibiotic Blood Levels, Parasitology, physicians to undertake bacteriological work in and Virology were established in the Infectious the department. In 1925, just before the end of Dr. Diseases Unit. All these had close relationships Wright’s tenure as chief, the offi cial position of with the Bacteriology Laboratory as well. In 1989 Bacteriologist was established in the Department the activities of the Bacteriology/Clinical Micro- of Pathology, and this was held in succession by biology Laboratory were once again returned to Drs. George Lawson and Merrill King over the Pathology under Clinical Pathology. With the next several years. In 1930 Dr. Louis Dienes was consolidation of laboratories in the early 1990s, appointed Bacteriologist, a position he held until the activities in all these specialized infectious 1952. Under his direction the fi eld of bacteriology diseases laboratories were ultimately incorpo- began to fl ourish and the number of diagnostic rated under Bacteriology to make it a true Clini- studies carried out in the Clinical Bacteriology cal Microbiology Laboratory. Before this, how- Laboratory increased dramatically. After Dr. ever, Dr. Kunz had retired in 1982 and Dr. Mary Dienes’s retirement in 1952, Th omas Fite Paine Jane Ferraro had been appointed his successor. Jr. was appointed Chief of Bacteriology, and a She has maintained leadership of the Clini- separate department was established, beginning cal Microbiology Laboratories until the present a nearly four-decade period (1952–1989) in which time. Th us, in a period of just less than a century, clinical microbiological studies were performed Clinical Microbiology at the MGH evolved from outside the Pathology department. Upon Paine’s a nascent discipline in Pathology to a separate

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Department of Bacteriology, which ultimately leishmaniasis. Dr. Wright’s monograph entitled became a full-fl edged Division of Clinical Micro- “Th e Biology of the Microorganism of Acti- biology, and was returned to its original roots in nomycosis” led to his receipt of the Samuel D. Pathology, where it now resides as a fl ourishing Gross Prize (53). During Dr. Wright’s tenure, lab- clinical, research, and teaching operation. oratory testing for infectious diseases was estab- During the fi rst hundred years of Bacteriol- lished in Pathology, although the number of tests ogy/Clinical Microbiology at the MGH, the lab- carried out was relatively modest by today’s stan- oratory occupied a number of diff erent venues. It dards. Initial tests involved studies of blood and began its existence under Dr. Wright in the Allen urine but were expanded to include cerebrospi- Street Building. By the 1930s bacteriology was nal fl uid before Wright retired (1). After visiting carried out in a building adjacent to the Allen a laboratory in Germany, Wright commissioned Street Building (see below). In the early 1950s the the building of the fi rst water-jacketed incuba- laboratory was relocated to the fourth fl oor of tor in the United States (fi gure 21.1), which was the Domestic Building, where it remained until built by a Boston coppersmith named Peter Gray. it was moved to a temporary building erected Th is gas-powered incubator served to provide the on the lawn in front of the Bulfi nch Building to appropriate environment for early bacterial cul- enable the destruction of the Domestic Building tures at the institution. It was placed on display and construction of the Gray Building. In 1968 in the MGH Ether Dome after its utility in the the laboratory moved into new quarters on the Bacteriology Laboratory ended. fourth fl oor of the Gray Building, which had A review of the annual reports of the Depart- been made possible through a bequest from the ment of Pathology to the MGH Board of Trustees estate of Mrs. Stephen S. Fitzgerald, in memory provides an interesting insight into the microbi- of her father, Francis Blake. Blake was the inven- ology activities carried out in Pathology during tor of the Blake transmitter, which was developed the Wright era (1). As early as 1898, Dr. Wright for the Bell Telephone Company, and was once a reported that Pathology was involved in the Trustee of the MGH. Th e Francis Blake Bacteri- “application of bacterial culture tests in cases of ology Laboratories provided much-needed space suspected diphtheria in the hospital wards” and and facilities for the growing activities of the Bac- “the testing of tissues and fl uids for the presence of teriology/Microbiology Laboratory. the bacillus of tuberculosis.” His 1899 report notes that Pathology had begun “examination of spu- The Wright Era tum for tuberculosis bacillus” and also observes Dr. James Homer Wright (chapter 4) served as that the laboratory was carrying out examinations Chief of the Department of Pathology at MGH of blood for the “malaria parasite.” He then went from 1896 until 1926. In the early 1900s, Dr. on to describe an interesting test for typhoid fever Wright authored sections titled “Actinomyco- that was employed in 1899 but is not mentioned sis” and “Diseases Due to Vegetable Parasites in subsequent reports: “In typhoid fever the blood Other Th an Bacteria” for Dr. William Osler’s test is of greatest value confi rming the diagnosis. textbook Modern Medicine, evidence of his grow- Th e test consisted of a drop of blood from earlobe ing interest in infectious diseases (28). In 1903 or fi nger-tip mixed with a few drops of bouillon he reported on the discovery of a protozoan in containing living typhoid bacilli, which caused the a tropical ulcer, but, although this was an inde- bacilli to gather together as clumps or groups.” He pendent observation, Dr. Wright was six months also noted that the laboratory was doing throat later than Charles Donovan in what turned out cultures for diphtheria and stated that the labora- to be the discovery of the causative organism of tory was beginning to study methods to cultivate

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Figure 21.1 James Homer Wright (left) in his laboratory with the ﬁ rst water-jacketed incubator (far right) in the United States. Th e person standing at the bench at the back of the laboratory is most likely Louis Brown.

bacteria in the absence of oxygen “with special ref- prolifi c in this area, publishing eight articles on erence to the bacillus of tetanus or ‘lockjaw’ ”; and the subject in 1897 and 1898 alone. He went on to in 1900 he published a paper describing a rather serve on the board of public health and published crude but eff ective method for culturing anaero- on inoculation against typhoid fever. Dr. Steele bic bacteria (57). It was at this time that he also served as Assistant in Clinical Bacteriology from began his own personal studies of actinomycosis 1910 through 1926. He had trained at Boston (fi gure 21.2). Th ese were remarkably pioneering City Hospital and at MGH. In MGH Pathology eff orts in early bacteriology, especially given the he played a major role in the therapeutic use of fact that they took place less than a quarter cen- killed bacteria. tury after the establishment of the microbial basis In 1907 the Pathology Department began to for bacterial infections by Robert Koch and Louis treat “certain bacterial diseases by subcutane- Pasteur. ous injections of killed cultures of the infecting Wright’s diagnostic eff orts and research studies organism” (1). Hundreds of patients received in bacteriology were supported by several other this therapy on the wards and in two separate physicians working in the Pathology department, rooms in the Pathology Department. Indeed, including Drs. Mark Wyman Richardson and 500 patients were treated in 1908, with “very Albert E. Steele. Richardson did pioneering work satisfactory results.” It should be noted that this on methods for diagnosing typhoid fever and was type of therapy was carried out in Pathology for

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appointed Bacteriologist in his place, but he left in 1930 after a brief tenure to enter a career in ophthalmology, which led to the appointment of Dr. Louis Dienes (53). The Dienes Era Dr. Louis Dienes (fi gure 21.3) was appointed Bacteriologist in the Department of Pathology in 1930, a position he held until his retirement in 1952 (53). Dienes was born in Tokay, Hungary, in 1885 and received a medical degree from the University of Budapest in 1908. In 1913 he began to work in bacteriology research in the laboratory Figure 21.2 “Human Actinomycosis smear preparation of Professor Fred Neufeld at the Robert Koch from a ‘granule’ showing the branching fi laments of the Institute in Berlin. He then served as an Army microorganism’ ” by James Homer Wright fi eld bacteriologist in Poland and Ukraine dur- (photograph by Louis S. Brown) ing World War I, which provided him with a vast experience in outbreaks of cholera, typhoid, and the next several decades (although in decreas- typhus fever. He developed a particularly valu- ing numbers), but no formal evaluation of the able collaboration with Professor Edmund Weil its results were ever given in the reports. In 1911 in the study of epidemic typhus fever and his the laboratory began studies of the urinary excre- work on the Weil-Felix reaction began to focus tion of Salvarsan (discovered by Paul Ehrlich in his interest on the immune response in infec- Germany only two years earlier) and began work- tious diseases. In 1922 he went to the von Ruck ing on a modifi ed serological test for syphilis. Research Laboratory at a tuberculosis sanitarium By 1912 Dr. Wright’s laboratories were carrying in Asheville, North Carolina, where he began out more than 2,000 “bacterial evaluations for to study the immune response in tuberculosis. diphtheria, typhoid fever and other diseases” and Th rough this research he was able to characterize had inoculated 300 guinea pigs for tuberculosis. the histological features of the tuberculin reac- Th e serological testing for syphilis had begun in tion and noted that these are distinctive reactions earnest, and 2,262 Wassermann tests were per- that diff er from other types of infl ammation and formed in the laboratory that year. Th e number from immunologic reactions mediated by circu- of bacterial examinations carried out in the labo- lating antibodies—a phenomenon we now know ratory remained essentially stable until near the as delayed-type hypersensitivity (47). He also end of Dr. Wright’s tenure, but the number of noted that delayed-type hypersensitivity can be serological tests for syphilis had increased almost elucidated with other nonbacterial antigens and eightfold, to 16,291 in 1925. In 1926 the offi cial is not unique to infection with the tubercle bacil- position of Bacteriologist was established in the lus. His work also paved the way for using an Department of Pathology, and this was fi lled in adjuvant to enhance the development of delayed- rapid succession by Dr. George Lawson and then type hypersensitivity. by Dr. Merrill King. Lawson was appointed Bac- In 1930 Dr. Dienes moved to Boston to teriologist in 1927, but in 1928 he left to accept work with Hans Zinsser in the Department a position as Associate Professor of Bacteriology of Bacteriology and Immunology at Harvard at the University of Louisville (53). Dr. King was Medical School (HMS), and he was appointed

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horses’ wagons used for transport of patients to the hospital. Th e small building on Allen Street separated these important facilities from the rest of the hospital. Included among them were the Pathology Laboratories, the autopsy room, and the “cold room,” where cadavers were held. A large amphitheater overlooked the autopsy area. Also present were the Bacteriology media-mak- ing room, with its stoves and autoclaves, and the “animal farm,” which was a vital research facil- ity. Despite these limitations, Dienes was able to carry out his important research and to oversee the growing clinical activities of the Bacteriology Figure 21.3 Louis Dienes Laboratory. Dr. Dienes continued the bacterial examina- Bacteriologist at the MGH, where he contin- tions for diphtheria, typhoid, and other organ- ued his studies of delayed-type hypersensitiv- isms started by Dr. Wright. He also performed ity in close collaboration with Tracy B. Mallory the serological tests for syphilis, which continued until the mid-1930s (chapter 23). At that point to grow in volume (1). In 1928 the laboratory he changed research interests and began studying switched from the Wassermann to the Hinton the so-called L-forms derived from Streptobacillus test after it was developed by William A. Hinton, moniliformis. His work defi ned the stages that led a professor at HMS who had worked in MGH to the development of L-forms from the bacillary Pathology in the early 1920s (chapter 3). Th is test form of S. moniliformis, and he demonstrated for syphilis was thought to be more accurate than the spontaneous development of L-forms from the earlier Wassermann examination. In 1933 the other pleomorphic organisms (7). He was able to laboratory began typing pneumococci, and the show that penicillin and glycine could be used to number of bacterial examinations carried out by induce the development of L-forms of the spe- the laboratory increased to 5,160; 20,773 Hinton cies in which they did not occur spontaneously tests were also performed in 1933. Although the and ultimately demonstrated through electron number of patients treated with bacterial vac- microscopy that L-forms lack a complete cell cines in Pathology persisted, the actual num- wall (8). His work with L-forms led to work with ber of patients receiving this therapy continued a morphologically similar pleuropneumonia-like to decrease, and by 1940 only 30 patients were organism, which he initially isolated from the treated with vaccines in Pathology. Th at year the genital tract of a patient in the late 1930s. Th ese number of bacterial cultures jumped dramati- organisms, which are now known to be myco- cally, to 17,292, a direct result of the discovery of plasmas, were fi rst isolated by Dr. Dienes in pure the sulfonamides and the beginning of the antibi- culture from an abscess of a Bartholin’s gland otic era. Guinea pig inoculations for tuberculosis cyst. totaled 1,056, and 831 pneumococcal typing tests During Dr. Dienes’s early years at the hos- and 30,150 Hinton tests for syphilis were carried pital, offi ce and laboratory space was primitive out in the laboratory. In 1951, just a year before by today’s standards. Th e building housing the Dr. Dienes retired, it was noted that the number Bacteriology Laboratory was over 100 years old of bacterial cultures had remained stable over the and had formerly been the stable housing the preceding decade, at just over 17,000 (1).

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Louis Dienes retired as head of the Bacte- riology Laboratory in 1952 but continued his research in a small laboratory at the MGH where, with the technical assistance of Sarabelle Madoff Annenberg, he collaborated with members of the Arthritis (now Rheumatology) Unit until 1970. He died on January 31, 1974. In 1976 his daughter, Dr. Priscilla D. Taft, Director of MGH Cytopathology, was among those present for the unveiling of the plaque dedicating the Louis Dienes Conference Room in the Francis Blake Bacteriology Laboratories. The Kunz Era Upon the retirement of Louis Dienes, major changes occurred in the Bacteriology Laboratory. Th omas Fite Paine Jr. was named Chief of the Bacteriology Laboratory in 1952, and the Bacteri- ology Laboratory became a separate, independent department, severing its direct ties with Pathol- Figure 21.4 Lawrence J. Kunz ogy (11). Paine’s tenure, however, was short, and he left within a year to take a position at Van- Laboratories on Gray 4 (now Gray 5) served derbilt University. At that point Dr. Lawrence a number of important functions. It provided J. Kunz, who had been serving as an Assistant the space and facilities for rapid expansion of Bacteriologist at the MGH, assumed respon- the laboratory and, because of its proximity to sibility for the laboratory; in 1956 he was made the Infectious Diseases Unit, it allowed many permanent Chief of the Department of Bacteri- fruitful collaborations between Bacteriology and ology (fi gure 21.4). Kunz was born in Brooklyn, Infectious Diseases. Before the movement of the New York, where he graduated from Brooklyn laboratories to Gray 4, the cramped space in the Preparatory School. He attended and graduated old Domestic Building and its relative isolation from St. Peter’s College in New Jersey. Following precluded many of the advantages realized after service in the 15th Medical Corps in Italy during the move to the Gray Building. In the early 1960s World War II, he earned a Ph.D. in bacteriol- there was very little if any contact between mem- ogy from Harvard University, working with the bers of the MGH house staff and the laboratory. Nobel laureate Dr. John Enders. Infectious Diseases was in its infancy, Dr. Morton Under Kunz’s direction, the Bacteriology Lab- Swartz having been appointed chief of the nascent oratory was ushered into the modern era. By the unit in 1956, which in the early 1960s consisted late 1950s the volume of specimens for routine of Drs. Swartz, Frank Austen, Paul Black, and, bacterial cultures, fungal cultures, mycobacte- subsequently, Arnold Weinberg. Drs. Kunz and rial cultures, and serology had skyrocketed as Weinberg, who was also from Brooklyn, quickly medicine entered the expanding antibiotic era, became friends and close colleagues; Weinberg which allowed for specifi c treatment of infec- joked that although he didn’t know Kunz as a tions with known etiology. Th e relocation of the youth, he strongly suspected that Kunz was one laboratory to the new Francis Blake Bacteriology of the boys that used to throw stones at him as

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he walked through his neighborhood on his way Disease Control (CDC) on the taxonomy of the to school! so-called paracolon bacilli, which includes many Many diagnostic activities were carried out or of the currently recognized species of Enterobacte- initiated outside the laboratory. In 1955 there were riaceae (13). He also was among the fi rst to make incubators on Burnham 6, on Bulfi nch 2, and eff ective utilization of the Bacteriology Labora- in the Emergency Ward so that house staff and tory for the identifi cation of nosocomial infec- physicians could plate their own cultures when tions. Th us, with Dr. Orjan T. C. Ouchterlony, a the Bacteriology Laboratory was not open nights Swedish physician and bacteriologist who was a and weekends. A memorandum from Dr. Kunz visiting professor at MGH in 1954, he and his col- in 1956 reminded the house staff that “plated out leagues discovered that brewers yeast contained cultures which were submitted to the laboratory three diff erent types of Salmonella as contami- must be streaked out on the surface of the agar nants that were causing infections in hospitalized with an inoculating needle. Specimens streaked patients (25). One of the most intriguing clinical out with a swab are unsatisfactory.” Th e laborato- investigations involved the solving of an outbreak ries in the Emergency Ward and on all the major of nosocomial infections at the MGH with a rare clinical units provided materials for house staff species of Salmonella, Salmonella cubana, which to carry out their own Gram stains, buff y coat was ultimately traced to carmine red dye that was smears, routine urinalysis, and blood counts. As used in the 1960s as a measure of gastrointesti- a result, members of the medical house staff in nal motility (27). Th e detective work carried out particular were well trained in carrying out and by Kunz, his colleague Dr. David Lang, and a interpreting Gram stains of sputum, urine, and laboratory technologist, Harriet Provine, served other specimens. as the basis for a number of scientifi c papers and In 1955 Massachusetts experienced a major generated a New Yorker article by Berton Roue- polio epidemic. Because of the large number ché entitled “Th e Santa Claus Culture.” With of patients with respiratory diffi culties and the closer collaboration between the Bacteriology shortage of professional personnel on the wards, Laboratory and members of the Infectious Dis- as well as the need for repeated bacteriological eases Unit, a number of important clinical inves- cultures, particularly of the respiratory tract, a tigations were carried out and published. Th ese special section of the Bacteriology Laboratory included major studies of anaerobic infections, was temporarily set up on White 9 in the charge extrarespiratory streptococcal infections, salmo- of a second-year medical student, John Living- nellosis, listeriosis, endocarditis due to group D stone. Fortunately, Livingstone had been work- streptococci or enterococci, and infections due to ing in the Bacteriology Laboratory during the Acinetobacter calcoaceticus (2, 9, 17). summer, and that made it possible for him to In 1968, while serving as a research fellow in take charge of this special unit, which monitored Infectious Diseases at the MGH, Dr. Robert C. the fl ora of patients who were at continual risk Moellering Jr. began a series of collaborations of contracting bacterial infections. Livingstone’s with Dr. Kunz in the Bacteriology Laboratory. work not only provided immense benefi t to the By using novel sealed agar plates provided by patients stricken with polio, but it resulted in a Dr. Kunz, Dr. Moellering was able to carry out 1957 publication in the New England Journal of a number of cultures under jungle conditions in Medicine (29). the Solomon Islands (34). He used this method Dr. Kunz began a number of research studies to obtain soil cultures from the environment as in the laboratory, beginning with work in collab- well as wound and stool cultures from the natives oration with William Ewing at the Centers for in an area on Malaita, where antibiotics had not

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previously been used clinically. From these cul- errors in susceptibility test results or organism tures Dr. Moellering and his colleagues isolated identifi cation, based on “sensi-ID discrepan- bacteria with transferable resistance to strepto- cies” (37). Th ese activities, which were unique to mycin and tetracycline, thus providing the fi rst MGH at the time, have now become routine or evidence that resistance genes existed in nature mandated in today’s microbiology laboratories. independent from the clinical use of antimicro- Clinical microbiology laboratories are among bial agents (14). Dr. Moellering also provided the few laboratories analyzing clinical specimens soil samples to the Mycobacteriology section of that have proven diffi cult to automate. Appro- the MGH Bacteriology Laboratory, from which priate isolation and identifi cation of bacteria, were isolated a series of “atypical” mycobacteria viruses, fungi, and other pathogens still require that probably provided the source of positive individual, subjective, technical judgment. Dr. skin tests for atypical mycobacteria among the Kunz, however, was among the fi rst to realize the Solomon Islanders. Th is work served to suggest potential of the computer for the accurate collec- strongly that exposure to “atypical” mycobacteria tion and analyzing of data generated in the Bac- does not provide protective immunity to tuber- teriology Laboratory and for transmission of the culosis, which had recently been introduced into data to the clinicians caring for infected patients. the Solomon Islands (32, 42). He collaborated with members of the Labora- Upon completion of his clinical and research tory of Computer Science at MGH, including fellowship in Infectious Diseases, Dr. Moellering James Poitras and Dr. G. Octo Barnett, to create joined the full-time staff in the Infectious Diseases a system that was uniquely suited to the needs Unit and established an independent research of the hospital (26, 37). Although a number of laboratory. He then received a small hospital sti- commercial systems for microbiology laboratory pend to serve as a formal Consultant to the Bac- computerization have been developed in sub- teriology Laboratory and collaborated with Dr. sequent years, none has proven more fl exible, Kunz in a number of projects relating to anti- sophisticated, and user-friendly than the original microbial susceptibility testing and laboratory program developed by Kunz and his colleagues. automation. Th ey developed a novel mechani- Not only did the system provide a valuable tool cal method for inoculating plated media used for analyzing large amounts of data and provid- in antibiotic susceptibility tests (24) and began ing suitable reports on susceptibility patterns, evaluation of a computer-associated electronic frequency of isolation of various pathogens from zone analyzer, which fed data on antimicrobial specifi c body sites, and the like, but it also served susceptibility testing directly into the laboratory to eliminate many of the transcription errors that computer (37). Th is proved to be a powerful tool occur when massive amounts of data are analyzed for evaluating antimicrobial susceptibilities of a and transmitted manually. large number of organisms in the laboratory and During this period Dr. Barbara K. Watson was yielded invaluable data for evaluation of new recruited to the laboratory, joining Dr. Th eodore antimicrobial agents such as tobramycin and F. Medrek in a research investigation on strepto- amikacin (35), for monitoring the development cocci. Th ey carried out studies relating to strepto- of antibiotic resistance, and for providing annual coccal taxonomy in the Bacteriology Laboratory, summaries of institutional-based data to guide and both collaborated with Moellering and other initial empiric therapy (4, 36, 38, 39). By feeding members of the Infectious Diseases Unit in stud- data directly from susceptibility test results into ies relating to their clinical or research activities. the computer, it was also possible to develop a Particularly useful among Dr. Watson’s studies real-time quality assurance system that identifi ed were a number of projects defi ning more accurate

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ways to identify streptococci to the species level (9, 54, 55). Dr. Watson retired in 1975. Eventually, Dr. Kathryn L. Ruoff , who joined the Bacteri- ology Laboratory in 1981 as an Assistant Bacte- riologist, continued the research work on streptococci and produced some particularly valuable results on the pathologic signifi cance of S. bovis and related species (46). Dr. Ruoff received her Ph.D. degree from Cornell University in 1975 and early in her professional career taught microbiology at Cornell and at Wheaton College in Massachusetts. She contributed greatly to teaching her discipline both in the MGH laboratory and at HMS, where she was promoted to Associ- ate Professor of Pathology in 1997. In 2002 she left MGH to assume a position at Dartmouth- Hitchcock Medical Center and was appointed Figure 21.5 Robert C. Moellering Jr. Associate Director of Microbiology in 2007. pharmacokinetic parameters and dosage recom- The Boutique Infectious Diseases mendations that had previously not been avail- Laboratories able for drugs such as vancomycin (16, 23, 33). In Because of a growing need for specialized clinical the mid-1970s the research laboratory began to laboratory diagnostics beyond those of bacteriol- serve as a clinical laboratory for antibiotic assays, ogy and mycology, the Department of Medicine’s and fl uorescent polarization enzyme immu- Infectious Diseases Unit established three small, noassays eventually became available commer- independent clinical laboratories during the cially and replaced the labor-intensive bioassays. 1970s. Th ese laboratories were among a number Shortly after Dr. Moellering’s departure in 1981 to of boutique laboratories that had sprung up over become Chair of the Department of Medicine at the years at the MGH and performed tests for New England Deaconess Hospital, the activities Parasitology, Antibiotic Blood Levels, and Virol- of his former Antibiotic Blood Level Laboratory ogy; the latter two arose from active research were consolidated into the Microbiology Labora- programs that found themselves also providing tory. In December 1991 these assays were trans- necessary but unfunded clinical services. ferred to the Chemistry Laboratory, allowing In the early 1970s Dr. Robert Moellering (fi g- for seven-day service, more automated enzyme ure 21.5) and Christine Wennersten began to carry immunoassay testing, and rapid reporting. out assays for concentrations of antimicrobial Th e Parasitology Laboratory was developed in agents in patients’ serum in Moellering’s research the Infectious Diseases Unit at approximately the laboratory. Initially these laboratory-developed same time. Initially supervised by Drs. Gordon bioassays were used to measure serum levels of Moore, an infectious diseases fellow who had aminoglycosides (gentamicin, tobramycin, and obtained training in parasitology at the CDC, amikacin) and vancomycin and allowed for the and Dr. Morton Swartz, the laboratory was sub- study of appropriate dosage of aminoglycosides sequently overseen by Dr. John Wolfson. Barbara in specialized situations, such as children with Parkhurst and later Mary Ann Waldron served as extensive burns, and for the determination of the supervisors and major laboratory resources

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for this operation. Th ey assisted Dr. Wolfson in a series of annual lectures on the clinical and laboratory diagnosis of parasitic diseases, which were mandatory for infectious diseases fellows and off ered to Pathology residents. Th ick and thin smears for the laboratory diagnosis of malaria, one of the true medical emergencies in infectious diseases, were carried out by infectious diseases fellows during evenings, nights, and weekends, when the laboratory was not open. In 1991, following Dr. Wolfson’s untimely death, the Para- sitology Laboratory was incorporated into the Clinical Microbiology Laboratory, which allowed the eventual cross-training of technologists and incorporation of test results into the Microbi- ology Laboratory’s computer-generated patient reports. Dr. Anthony Mattia assisted in the direction of this section of the laboratory from 1993 to 1994. Th e need for a Diagnostic Virology Labora- tory at MGH was recognized in the early 1970s. Improvements in culture techniques for a variety of viruses, early studies on antiviral therapy Figure 21.6 Martin S. Hirsch at MGH and elsewhere, and curtailment of state and federal diagnostic services combined to application was approved, equipment was pur- emphasize the need for an in-house laboratory. chased, and the fi rst laboratory personnel were Until 1974, the few MGH diagnostic studies hired in late 1974 and early 1975. done were performed as part of therapeutic stud- During the fi rst year of operation, 771 speci- ies in herpes simplex encephalitis and varicella- mens were received for virus isolation, of which zoster virus infections and were performed in the 129 were positive, and 1,271 specimens were research laboratory of Dr. Martin Hirsch of the received for antibody studies. In 1977 the fi rst Infectious Diseases Unit. attempts at rapid diagnosis (immunofl uorescent Early in 1974 Dr. Hirsch (fi gure 21.6) initiated detection of herpes group viruses) were begun, discussions with Dr. Lawrence Kunz, Director and in 1979 chlamydia diagnosis was added to of the MGH Bacteriology Laboratory, regard- the tests off ered (10, 21, 51). Th e Virology Labo- ing a joint eff ort to begin diagnostic virology at ratory was incorporated into the Microbiology the hospital. Dr. Kunz generously off ered space Laboratory in 1991 and subsequently directed on Gray 5 for this purpose, and Drs. Hirsch and by Drs. Mary Jane Ferraro and Angela Caliendo Morton Swartz, Chief of the Infectious Dis- (see below). Dr. Hirsch, however, remained a val- eases Unit, prepared an application to the MGH ued consultant to Microbiology and continued Resource Allocation Board for approval of such a to participate in important decisions regarding laboratory, with the active support of many other implementation of more rapid nonculture meth- departments (e.g., Medicine, Pediatrics, and Sur- ods for viral diagnosis (31). gery) that would benefi t from such a service. Th is Each of these three laboratories in the

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Infectious Diseases Unit had close ties with the the direct Pathology appointments of several Bacteriology Laboratory, and in the early 1990s, key members from the Infectious Diseases Unit when administrative responsibility for the Micro- as Assistant Directors, starting with Dr. Angela biology Laboratory was returned to the Depart- Caliendo in 1994 and Dr. Eric Rosenberg in ment of Pathology, they were incorporated into 2002. In many ways this association has been a the clinical laboratory to form the modern-day model for collaborative eff orts between academic Clinical Microbiology Laboratory. Th e establish- departments of Pathology and Medicine in joint ment of these specialized clinical laboratories in research and training activities. the Infectious Diseases Unit and their ultimate Dr. Lawrence Kunz retired from his position consolidation into the Clinical Microbiology as Chief of the Clinical Microbiology Labora- Laboratory in Pathology marked the continua- tory in December 1982. In 1993, 11 years after his tion of a close and mutually benefi cial associa- retirement, Kunz was honored by having a newly tion between the laboratory and the Infectious classifi ed bacterial species, Helcococcus kunzii, Diseases Unit. Th e association began informally named after him (6) (fi gure 21.7). Appropriately, with collaborative studies between Bacteriology this organism had been discovered and charac- and the newly formed Infectious Diseases Unit in terized at the MGH by Dr. Kathryn Ruoff , who the 1960s, as noted earlier. It was codifi ed in 1973 was instrumental in its naming. Dr. Kunz died when Dr. Moellering was named a formal Con- on October 27, 1999. sultant in Bacteriology, a position he held until his departure in 1981. In 1982 Dr. Arnold Wein- The Present Era berg was named Consultant in Bacteriology and Dr. Mary Jane Ferraro took over direction of provided valuable assistance to the laboratory for MGH Microbiology upon Dr. Kunz’s retirement several years. Ultimately, this position was incor- in 1982. Dr. Ferraro received her Ph.D. in Medical porated more formally into the laboratory with Sciences–Microbiology from Boston University

Figure 21.7 Reception in honor of the newly classiﬁ ed bacterial species Helcococcus kunzii. Left to right: Mary Jane Ferraro, Lawrence J. Kunz, Kathryn L. Ruoﬀ , Arnold N. Weinberg, Katherine Kunz, Morton N. Swartz.

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School of Medicine, where she worked with Dr. provided a valuable early warning system for the Sidney Cooperband. She then completed stud- clinicians at the MGH and other local institu- ies leading to an M.P.H. and postdoctoral train- tions concerning possible seasonal outbreaks of ing at the Harvard School of Public Health, respiratory virus infections. which included internships at the Massachusetts Th e appointment of Angela Caliendo as Assis- Department of Public Health Laboratories and tant Microbiology Laboratory Director in 1994 MGH. She was recruited to the MGH in 1976 as marked the further introduction of molecular a junior staff member in Medicine and Assistant diagnostic methods in Microbiology, particularly Director in the Bacteriology Laboratory. In the those utilizing polymerase chain reaction (PCR). laboratory she began a number of research stud- Dr. Caliendo was exceptionally well trained to ies relating to automated, rapid, and cost-eff ec- develop this new service, having received her tive diagnostic methods, antimicrobial suscepti- Ph.D. in biochemistry in 1983 from Case West- bility testing, and other applications of clinical ern Reserve University, where she continued microbiology (3, 12, 18, 20, 22, 43, 45, 52). Her studies leading to her M.D. in 1987. Following work has led to setting worldwide standards and her residency in internal medicine at Brigham guidelines for susceptibility testing of bacteria and Women’s Hospital, she completed a fellow- and other microorganisms (48, 49). She became ship in infectious diseases at the MGH. Her ini- Professor of Pathology in 2004 and Professor of tial eff orts led to the development of a “home- Medicine in 2006, which places her among a grown” system for identifying herpes simplex handful of women ever to have held dual HMS virus in the cerebrospinal fl uid (CSF) of patients professorships. with meningitis and encephalitis. Following this, Under Dr. Ferraro’s direction there was a rapid the laboratory initiated HIV-1 viral load testing expansion in laboratory services, including, as and made the test available for patient care long noted earlier, the incorporation of parasitology before it was FDA-cleared. Th is revolutionized and virology operations in the Clinical Micro- the care of HIV-infected patients at MGH and biology Laboratory and the implementation provided the support for many seminal clinical of a night shift, thereby creating the fi rst 24/7 studies of HIV in collaboration with members microbiology service operation and thus elimi- of the Infectious Diseases Unit (30, 40, 44, 50), nating the need for house staff and infectious including the defi nitive description of acute HIV diseases fellows to inoculate important cultures infection as a mononucleosis-like syndrome by and perform Gram stains during off -hours. New Dr. Eric Rosenberg during his infectious diseases approaches to nonculture-based laboratory test- fellowship. During the mid-1990s, Drs. Cali- ing were implemented, including direct antigen endo, Ferraro, and their colleagues also studied testing for some bacterial and fungal pathogens as or incorporated a number of molecular diagnos- well as nucleic acid–based tests for the detection tics, including PCR for cytomegalovirus (19) and of Neisseria gonorrhoeae, Chlamydia trachomatis, Pneumocystis carinii (5), and molecular tests for and Mycobacterium tuberculosis (56). A same- atypical respiratory pathogens such as Mycoplasma day cytomegalovirus (CMV) antigenemia test pneumoniae (the genera originally studied by Dr. replaced slower CMV blood cultures (31), and Louis Dienes), and Chlamydia pneumoniae. Dr. respiratory virus cultures were discontinued in Caliendo left MGH in 1999 to become Medical favor of rapid direct fl uorescent antibody (DFA) Director of Microbiology and Molecular Diag- testing for respiratory viruses. Moreover, by post- nostics at Emory University in Atlanta, and she ing the summaries of the DFA results of tests for was subsequently named Vice Chair for Clini- respiratory viruses in real time, the laboratory cal Pathology in the Department of Pathology

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Figure 21.8 MGH Microbiology, 2010. Left to right: JiYeon Kim (fellow), Eric S. Rosenberg, Mary Jane Ferraro, John A. Branda.

and Laboratory Medicine. Shortly after, Dr. In addition to its diagnostic and research James Versalovic, who had just completed clini- activities, the Clinical Microbiology Laboratory cal pathology residency training at MGH, joined continues its strong tradition of education. Sev- the Microbiology Laboratory as an Assistant eral members of the laboratory staff devote sig- Director with responsibility for molecular diag- nifi cant eff ort to teaching of medical students nostics. Dr. Versalovic left MGH in 2001 for in the clinical laboratory. Of particular note has Baylor University College of Medicine, where he been Dr. John Branda; an HMS graduate, he became Chairman of the Pathology Department was appointed Assistant Director of Microbiol- at Texas Children’s Hospital in 2009. In 2002 Dr. ogy in 2004 after fi nishing his residency in clini- Eric Rosenberg became Associate Director of the cal pathology at MGH, and he has been closely Microbiology Laboratories with responsibility involved in the teaching of microbiology at HMS. for the molecular microbiology section. A 1991 Pathology residents and infectious diseases fellows graduate of Mount Sinai School of Medicine, have regular rotations in the laboratory, where Dr. Rosenberg trained in internal medicine at they receive intensive immersion in all aspects of the University of North Carolina and completed clinical microbiology. Regularly scheduled “plate his infectious diseases fellowship at the MGH. rounds” are one of a number of joint teaching While still maintaining his research and clinical eff orts of the Clinical Microbiology Laboratory activities in infectious diseases, he increased the faculty and the Infectious Diseases Unit. In 2010 test menu to include PCR-based viral load tests a formal fellowship in clinical microbiology was for hepatitis viruses and genotyping. started; it was approved by the ACGME in 2011.

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In addition, medical or pediatric residents and References infectious diseases fellows and attendings regu- 1. Annual Reports of the Department of Pathol- larly stop by the laboratory to check on culture ogy to the Board of Trustees of the Massachusetts results and for informal training and consultation. General Hospital, 1895–1951. Clafi n Library and Conclusion MGH Archives. 2. Bornstein DL, Weinberg AN, Swartz MN, Kunz Clinical microbiology is now a full-service labora- LJ. Anaerobic infections. A review of current tory, with several faculty directors, a microbiology experience. Medicine 43:207–232, 1964. fellow (fi gure 21.8), and approximately 70 employ- 3. Branda JA, Lin TYD, Rosenberg ES, Halpern EF, ees, that processes over 500,000 specimens per Ferraro MJ. A rational approach to the stool ova year. Included among these many tests are occa- and parasite examination. Clin Infect Dis 42:972– sional requests for studies of the same type that 978, 2006. James Homer Wright performed to assist in the 4. Calderwood SB, Wennersten C, Moellering RC diagnosis of diphtheria, tetanus, malaria, typhoid Jr., Kunz LJ, Krogstad DJ. Resistance to six ami- noglycosidic aminocyclitol antibiotics among fever, and tuberculosis. Th ese studies, however, enterococci. Prevalence, evolution and relation- are overwhelmed by a plethora of new tests that ship to synergism with penicillin. Antimicrob have been brought online in recent years. Agents Chemother 12:401–405, 1977. Th e microbiology laboratories, over the more 5. Caliendo AM, Hewitt PL, Allega JM, Keen A, than 100 years of their existence, witnessed the Ruoff KL, Ferraro MJ. Performance of a PCR fi rst applications at the MGH of the discoveries assay for detection of Pneumocystis carinii from of Pasteur, Koch, and others—research that pro- respiratory specimens. J Clin Microbiol 36:979– vided the fi rst rational basis for the diagnosis and 982, 1998. ultimate treatment of infectious diseases. Th ey 6. Collins MD, Facklam RR, Rodrigues UM, Ruoff participated, several generations later, when the KL. Phylogenetic analysis of some Aerococcus- discovery and clinical application of the sulfon- like organisms from clinical sources. Description amides and penicillin ushered in the golden age of Helcococcus kunzii gen. nov., sp. nov. Int J Syst of the management of infectious diseases, mak- Bacteriol 43:425–429, 1993. 7. Dienes L, Edsall G. Observations on the L-organ- ing the activities of the clinical microbiology lab- ism of Klieneberger. Proc Soc Exp Biol Med oratory a key player in the battle against severe 36:740–744, 1937. infections. Th e last half of the twentieth century 8. Dienes L, Bullivant S. Comparison of the mor- and the dawn of the twenty-fi rst have seen strik- phology of PPLO and L-forms of bacteria with ing improvements in therapy for fungal and viral light and electron microscopy. Ann NY Acad Sci infections as well—all the more important as 143:719–733, 1967. modern medicine and the HIV virus have mark- 9. Duma RS, Weinberg AN, Medrek TF, Kunz LJ. edly increased the numbers of immunocompro- Streptococcal infections. Medicine 48:87–127, mised patients with a wide variety of infections 1969. unimaginable by James Homer Wright and his 10. Eron L, Kosinski K, Hirsch MS. Hepatitis in an colleagues. Th roughout all these changes, the adult caused by herpes simplex type 1. Gastroen- MGH microbiological resources have adapted to terology 71:500–504, 1976. meet each new challenge. Th e battle is far from 11. Faxon NW. Th e Massachusetts General Hospital, 1935–1955. Cambridge: Harvard University Press, over, but it is certain that the activities of MGH 1959. Microbiology will continue to evolve and provide 12. Ferraro MJB, Edelblut MA, Kunz LJ. Accurate benefi t for the patients at the MGH and else- automated identifi cation of selected Enterobacteri- where well into the future. aceae at 4 hours. J Clin Microbiol 13:151–157, 1981.

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