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Productivity of the Roll-Streak Method to Perform Anaerobic Bacteriology in the Routine Clinical Laboratory Thomas R Henry Ford Hospital Medical Journal Volume 24 | Number 2 Article 7 6-1976 Productivity of the roll-streak method to perform anaerobic bacteriology in the routine clinical laboratory Thomas R. Neblett Follow this and additional works at: https://scholarlycommons.henryford.com/hfhmedjournal Part of the Life Sciences Commons, Medical Specialties Commons, and the Public Health Commons Recommended Citation Neblett, Thomas R. (1976) "Productivity of the roll-streak method to perform anaerobic bacteriology in the routine clinical laboratory," Henry Ford Hospital Medical Journal : Vol. 24 : No. 2 , 103-118. Available at: https://scholarlycommons.henryford.com/hfhmedjournal/vol24/iss2/7 This Article is brought to you for free and open access by Henry Ford Health System Scholarly Commons. It has been accepted for inclusion in Henry Ford Hospital Medical Journal by an authorized editor of Henry Ford Health System Scholarly Commons. Henry Ford Hosp. Med. Journal Vol 24, No. 2, 1976 Productivity of the roll-streak method to perform anaerobic bacteriology in the routine clinical laboratory Thomas R. Neblett, PhD* OMPETENT recovery of anaerobic bac­ terica plus ability to provide, upon request, valid antimicrobial susceptibility data upon the isolates should be an essential service provided by the clinical microbiology labo­ ratory. Several methods are available to the laboratorian for this purpose.' ""Choice of Data presented are derived from one year anaerobe jars, roll-streak tubes or anaerobic of performing routine clinical anaerobic chamber will depend on: 1) number of beds bacteriology by the roll-streak tube method. in facility served, 2) amount of anaerobic 826 specimens from 1508 (54.8%) con­ bacteriologic activity required by the medi­ tained anaerobes. 162 specimens were poly­ cal specialties, and 3) the overall cost, space microbic (19.6%) and contained 528 strains and manpower considerations. Choice will of a total of 1051 isolated (50.2 %). 479 blood also depend upon the personal knowledge, specimens contained 525 strains of ana­ experience and preference of the clinical erobes. Propionibacterium, considered a microbiologist contaminant, occurred 209/210 times in blood as the sole isolate. Bacteroides oc­ In a clinical situation where research curred alone or polymicrobically 308 times upon anaerobic bacteria is not the prime and comprised 54.7% ofthe blood isolates mission the anaerobe bench should not be­ and 32.5% of the total anaerobic genera come the tail that wags the laboratory dog. Isolated. Clostridium, Peptococcus and Conversely, it should not be considered too Fusobacterium were the next most fre­ expensive, sophisticated or unusable be­ quently occurring anaerobes. The roll-streak cause laboratory personnel have not been method is a means to competent anaerobe adequately trained for it. The microbiology Isolation. Its apparatus is not expensive and laboratory which does not provide its clini­ permits use of routine laboratory incubation cian patrons with at least some simple effi­ equipment for anaerobic work. cient anaerobe culturing activity is not doing the entire job entrusted to it. The anaerobic bacteria may be defined broadly as a group of micro-organisms to which molecular oxygen is somewhat toxic. When their nutritional needs are met, their growth spans a moderately wide range of * Department of Pathology electrochemical conditions from human Address reprint requests to author at Henry Ford venous blood values (-El 80 mv) down nearly Hospital, 2799 West Grand Boulevard, Detroit Ml to the hydrogen electrode (-420 mv)'. Ana­ 48202 erobes tend to survive much better in their 103 Neblett natural habitat than in vitro, and the more vironment. Tally et al'* reported that some of nearly the microbiologist simulates in vitro the common anaerobes are able to survive the chemically reduced state of their (usu­ several hours' exposure to ambient condi­ ally) polymicrobic environment, the more tions, suggesting that factors other than mo- successful will be recovery from clinical lecular oxygen intoxication may be specimens. Violations of this principle can responsible for apparent anaerobe die-off. be responsible for a laboratory's failure to Swabs tend to shed only about 3-5% of achieve proficient anaerobe recovery. Ac­ organisms collected into their fibers,'" and a ceptance of improperly collected and han­ high death rate of collected bacteria will dled specimens, followed by inoculation result if desiccation is allowed to take place, onto oxygen-exposed commercially pre­ even if retained under oxygen-free gas." pared culture media in primary isolation Tissues, aspirated fluids and pus tend to be attempts, is an example of anaerobic superior to swab collected material for ana­ compromise. erobe recovery. A well-researched explanation of plastic Specimen collection toxicity to anaerobes has not been offered. Exposure of broth containing resazurin to Specimen selection for anaerobic culture certain plastics will cause a change to pink should be based upon suspected presence of color, indicatingoxidation ofthe fluid above oxygen-sensitive micro-organisms as princi­ Eh of -41 mv. Retention of molecular oxy­ ple disease-producing agents in locations gen within the pore structure ofthe polymer where they do not commonly exist in high has been hypothesized, since storage of normal population. Cervix, vagina and oral some of these products under nitrogen for cavity contents, feces and urines should not several weeks eliminates the oxidizing po­ be considered for anaerobic culture, except tential. The most promising commercial in special circumstances determined specimen collectors are made of glass. through physician-microbiologist consulta­ tion. Some type of specimen collection de­ Adequate skin de-germing to prevent con­ vice designed specifically to protect tamination of blood cultures with Pro- anaerobes should be employed, whether it pionlbacterium is essential.'^ A two-bottle be one prepared by the laboratory or pur­ system in which one remains unexposed to chased commercially. Several products traces of air is recommended. available commercially have excellent re­ covery potential. A feature common to most is their oxygen-free gas or liquid content. Several studies have been published dealing Methods with devices and methods of anaerobe spec­ Prior to 1966 Brewer jars were used in our imen collection.^"" laboratory for routine anaerobe isolation. Illuminating gas was used, and electrically- Contrary to a popular conception, all heated catalysts eliminated residual oxygen. specimens intended for the microbiology Routine plating media were used, and jars laboratory should not be stored under re­ usually were not opened for 48-72 hours. frigeration until delivery. Die-offof fastidious Later, we converted to safer Gaspaks. These anaerobes will be the penalty for chilling utilized a cold, palladium-coated alumina specimens intended for anaerobic recovery. pellet catalyst. A tube of methylene blue, Devices should not be used for anaerobe included in the jar, indicated by color loss collection which allow specimen material to thatthe Eh of the jar contents had reached -E be exposed to non-reduced fluids (such as 11 millivolts or lower. An improved technic certain transport media) or to plastics or was later adopted in which evacuation of Vi which do not provide a moist holding en­ atmosphere fol lowed by replacement with a 104 Roll streak method in anaerobic bacteriology mixture of oxygen-free 80% N2 -E 10% CO2 Late in 1970 this laboratory adopted the -E 10% H2 repeated 3 X was carried out. It roll-streak method of cultivating clinical produced anaerobic conditions more specimens under anaerobic conditions. Pre­ rapidly" and was less expensive than the sent anaerobic handling equipment also in­ Gaspak generator. Laboratory convenience cludes the evacuation-replacement jar dictated closing jars at day's end or when system, a flexible anaerobic glove box, and enough plates had been accumulated to fill a two thermal conductivity detector type gas- single jar, causing plates streaked for ana­ liquid chromatographic analyzers plus the erobic isolation to be left exposed to oxygen necessary gas handling regulators, valves for as much as several hours. and set-ups for the jars, roll-streakers, glove chamber and chromatographs. The decision to adopt the roll-streak was made after con­ Although improved between 1966 and ferring with other workers using the ana­ 1970, anaerobic procedures were still little erobic chamber method principally for different from routine culture methods. Thio­ research purposes. It was deemed inconve­ glycollate broth tubes were used to subcul­ nient, in a clinical laboratory situation, to ture from anaerobic blood collection bottles operate the airlock system necessary to and for routine anaerobic inoculation of maintain anaerobiasiseach timea specimen specimens such as spinal fluids. These were needed to be processed. The difficulty of never boiled to exhaust dissolved gases im­ others in maintaining gas-tight integrity with mediately prior to use. They were incubated rigid type plexiglass chambers suggested a aerobically, because reliance was placed potential maintenance problem. Since large entirely upon the reducing property of the numbers of jars were needed to handle the thioglycollate to provide anaerobic condi­ specimen load of a high volume laboratory, tions. Biochemical differentiations were car­ the decision became clear. ried out in ordinary tubed media containing acid-base indicators.
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