Thayer-Martin Research Articles

Rapid identification of Neisseria gonorrhoeae and Neisseria meningitidis by using enzymatic profiles

The enzymatic profiles of Neisseria gonorrhoeae, N. meningitidis, and related species were determined, using a total of 48 chromogenic substrates. Enzyme classes assayed for included glycosidases, aminopeptidases, phosphoamidases, proteases, lipases, esterases, and aryl sulfatase. A final test selection of 10 substrates, based upon their differential and reproducible characteristics, allowed the separation of N. gonorrhoeae and N. meningitidis from each other and from all species tested within 4 h after primary isolation on modified Thayer-Martin medium. The need for subculturing suspect colonies from modified Thayer-Martin medium to chocolate medium with a subsequent loss of 18 to 24 h of identification is eliminated.

Feasibility of Screening for Penicillinase-Producing Neisseria gonorrhoeae from Primary Culture Plates by Using a Rapid Microacidometric Test

We have shown that it is possible to screen for penicillinase-producing Neisseria gonorrhoeae directly from primary culture plates. Experiments involving the cocultivation of four genera of beta-lactamase-positive (beta-lac(+)) bacteria and a beta-lactamase-negative (beta-lac(-)) N. gonorrhoeae on modified Thayer-Martin medium indicated that suppressed or inhibited beta-lac(+) bacteria did not give false-positive results when isolated beta-lac(-) colonies of gonococci were tested. Colonies were assayed for beta-lac production by an adaptation of a microacidometric method in which bacteria could be tested before or after the addition of oxidase reagent.

Futility of routine culture of hysterectomy specimens for gonorrhea

Futility of routine culture of hysterectomy specimens for gonorrhea is briefly reported. Cultures were taken from the cervix of 200 consecutive hysterectomy specimens obtained from patients 20-86 years of age. The swab was immediately used for inoculation of Thayer-Martin medium under increased carbon dioxide. Oxidase testing of cultures was performed on 2 consecutive days. All cultures were negative for Neisseria gonorrhoeae indicating that the routine culture of hysterectomy specimens is not a productive procedure.

Imferon agar: improved medium for isolation of pathogenic Neisseria

Imferon, an iron-dextran complex, enhances the growth of Neisseria gonorrhoeae and N. meningitidis. The use of Imferon as a replacement for ferric nitrate, in a defined supplement for GC agar significantly increased the average colony sizes of both gonococci and meningococci. In comparison with Thayer-Martin medium, Imferon agar increased the speed and rate of isolation of gonococci from clinical specimens.

Simplified media for isolating Neisseria gonorrhoeae

Two simplified media for isolating Neisseria gonorrhoeae are described. They differ from the modified Thayer-Martin medium in the enrichment and antibiotics used for preparation. The enrichment for one, enrichment 4 medium, contains only four ingredients, and amphotericin B is used instead of nystatin. This medium is comparable to the modified Thayer-Martin medium for isolating N.gonorrhoeae. It is convenient to prepare and is only one-third the cost of Thayer Martin medium. It is a suitable alternative to the modified Thayer-Martin. The enrichment 5 medium is a hemoglobin-free version of enrichment 4 medium. It is somewhat more selective for contaminants but is also more inhibitory for N. gonorrhoeae.

Effect of a copper-containing intrauterine contraceptive device on Neisseria gonorrhoeae in vitro

The effects of a copper-containing intrauterine device on the growth of N. gonorrheae in vitro as determined on a typing medium developed by Kellogg et al are described. Plates were incubated for 24 or 48 hours in candle jars with an increased carbon dioxide atmosphere. N. gonorrheae were obtained from a patient with acute urethritis. The exudate was streaked on Thayer-Martin (VCN) medium. Copper wires .5 mm in diameter in 1-inch lengths copper plate cut into 1.5 cm discs and commercial copper-containing IUDs were placed on the typing medium which had been streaked with gonococci. All inhibited the growth of the organisms for 2 mm on each side. The solubilization of the copper has been estimated as 1 mcg of copper per day which is in the bactericidal range in vitro. Results suggest that copper-containing IUDs may play a useful role in preventing gonorrhea.

Screening Asymptomatic Male Patients For Gonorrhea

To the Editor.— The first few drops of urine, voided onto a dry swab and inoculated on Thayer-Martin medium, may be utilized to screen nonsymptomatic male patients for gonorrhea. It is now apparent that the female of the species is not the only hidden reservoir of gonorrhea. The male sexual contact of a person with gonococcal infection is also often an asymptomatic carrier, and routine screening of nonsymptomatic males admitted to a venereal disease clinic reveals 5% to 10% positive for Neisseria gonorrhoeae . The recommended procedure for securing an intraurethral specimen from the male without discharge has been use of a platinum loop or preferably a calcium-alginate-tipped, wire, urethrogenital swab. The loop or swab is then used to inoculate Thayer-Martin medium. 1 Either procedure generally requires presence of a physician, and the latter method is painful. An alternate procedure is urine culture of the sediment from a centrifuged specimen of

Pili on meningococci from primary cultures of nasopharyngeal carriers and cerebrospinal fluid of patients with acute disease.

The nasopharynx of known meningococcal carriers without signs of acute meningococcal disease as well as cerebrospinal fluid from patients with acute meningococcal disease were cultured on Thayer-Martin agar. Pili were observed in negatively stained preparations of over 80% cells from all primary cultures of both nasopharnyx and cerebrospinal fluids. Although pili were abundant on cells from all primary cultures, all pili were lost on serial subculture in the laboratory. This loss of pili from the cell surface on laboratory subculture was not accompanied by a concomitant loss of cell wall blebs.

New System for Cultivation of Neisseria gonorrhoeae

A modified Thayer-Martin medium, containing 2.0% agar, 0.25% dextrose, and 5.0 mug of trimethoprim per ml was compared to Thayer-Martin medium for growth of stock cultures of Neisseria gonorrhoeae and for isolation of N. gonorrhoeae from clinical specimens. Stock cultures and male urethral cultures grew equally well on both media, but the modified Thayer-Martin medium detected 10% more female patients positive for gonococci. A description is given of the use of a citric acid-sodium bicarbonate tablet for generating carbon dioxide in a sealed plastic bag containing a culture plate. Equivalent results were obtained on duplicate plates of 336 cultures and specimens when this procedure was compared with the standard candle jar extinction method of incubation. The tablet and plastic bag system has the advantages of economy, ease of use, and safety.

Comparison of Atmospheric Conditions for Culture of Clinical Specimens of Neisseria gonorrhoeae

We cultured 55 clinical specimens of Neisseria gonorrhoeae in the following atmospheric conditions: (i) 10% carbon dioxide in a CO 2 incubator; (ii) a candle extinction jar; (iii) an air convection incubator; and (iv) an anaerobic jar without added CO 2 . The number and size of colonies growing on modified Thayer-Martin medium were evaluated after incubation of cultures for 24 and 48 h at 36 C. After 24 h, the specimens from the candle extinction jar had the greatest number and size of colonies, but after 48 h growth was approximately equal for specimens from the candle jar and the CO 2 incubator. Only 19 of 55 specimens grew in the air convention incubator. None of 55 clinical specimens or of 10 laboratory strains grew anaerobically. Development of colonial morphology for colony types 1, 2, 3, and 4 was studied at 24 h on a base medium that contained no hemoglobin. The relative numbers of the four colony types in specimens were comparable after 24 h of incubation in any of the three atmospheric conditions under which growth occurred, but the different types were distinguished most readily when grown in the candle extinction jar.

Letters to the Editor

The information contributed by Dr. Ris regarding her success with vaginal cultures for N. gonorrhoeae is a welcome addition to the Academy's statement "Venereal Disease and the Adolescent," as are her comments on experience with Thayer-Martin medium. The incidence of the disease in the environment where Dr. Ris has conducted her studies has undoubtedly increased her preference for the mode of culturing she recommends. Meticulous attention to the details of culturing and the handling of the media en route to the incubator, as well as skillful laboratory interpretation, are of proven importance in recovering and identifying this elusive organism.

Zur IF-Diagnostik der Gonorrhö

The increase in gonorrhoea demands more thorough diagnostic measures. In 160 patients with urethritis and 60 female patients with various symptoms suggestive of gonorrhoea 3 different diagnostic methods (Gram staining, direct IF-test, cultural growth on Thayer-Martin medium) were compared. In the 160 male patients the 3 methods showed convergent results in 93%. The 10 cases yielding divergent results are discussed. In the 60 female patients the results were convergent in 86% from urethral smears, in 76% from cervical smears respectively. In 17 women gonorrhoea was diagnosed, 11 times with all 3 methods, 4 times with the culture and IF-test, twice with the IF-test alone. Comparing the results of the 3 methods, the IF-test has advantages both in use of time and equipment and has proved to be the most specific test in our experiments.

An Unclassified Gram-Negative Rod Isolated From the Pharynx on Thayer-Martin Medium (Selective Agar)

An Unclassified Gram-Negative Rod Isolated From the Pharynx on Thayer-Martin Medium (Selective Agar)

Comparison of Atmospheric Conditions for Culture of Clinical Specimens of Neisseria gonorrhoeae

We cultured 55 clinical specimens of Neisseria gonorrhoeae in the following atmospheric conditions: (i) 10% carbon dioxide in a CO(2) incubator; (ii) a candle extinction jar; (iii) an air convection incubator; and (iv) an anaerobic jar without added CO(2). The number and size of colonies growing on modified Thayer-Martin medium were evaluated after incubation of cultures for 24 and 48 h at 36 C. After 24 h, the specimens from the candle extinction jar had the greatest number and size of colonies, but after 48 h growth was approximately equal for specimens from the candle jar and the CO(2) incubator. Only 19 of 55 specimens grew in the air convention incubator. None of 55 clinical specimens or of 10 laboratory strains grew anaerobically. Development of colonial morphology for colony types 1, 2, 3, and 4 was studied at 24 h on a base medium that contained no hemoglobin. The relative numbers of the four colony types in specimens were comparable after 24 h of incubation in any of the three atmospheric conditions under which growth occurred, but the different types were distinguished most readily when grown in the candle extinction jar.

An unclassified gram-negative rod isolated from the pharynx on Thayer-Martin medium (selective agar).

An oxidase-positive, small gram-negative rod was isolated on Thayer-Martin medium (TM) inoculated with pharyngeal swabs obtained during surveys to detect Neisseria carriers. In one survey, this organism was isolated from 48% of the subjects, and 50 or more colonies were present on the majority of the primary isolation plates. Other characteristics of the organism, which has been given the provisional designation "TM-1," include: delayed production (2 to 10 days) of acid from glucose, formation of gas during nitrate reduction, and the frequent formation of "pits" in the agar surface. On TM, nonpitting colonies of TM-1 are morphologically similar to colonies of Neisseria gonorrhoeae and N. lactamica. Comparison of the characteristics of TM-1 strains with other similar fastidious gram-negative organisms encountered in clinical laboratories indicates that TM-1 is a distinct species. Further studies are required before proper taxonomic placement can be made.

Gonococcal Meningitis

In three cases of gonococcal meningitis, the organism was isolated from the spinal fluid by culture on Thayer-Martin medium. Two of these patients showed other manifestations of systemic gonococcemia. All three patients recovered completely following treatment.

An Unclassified Gram-Negative Rod Isolated From the Pharynx on Thayer-Martin Medium (Selective Agar)

An oxidase-positive, small gram-negative rod was isolated on Thayer-Martin medium (TM) inoculated with pharyngeal swabs obtained during surveys to detect Neisseria carriers. In one survey, this organism was isolated from 48% of the subjects, and 50 or more colonies were present on the majority of the primary isolation plates. Other characteristics of the organism, which has been given the provisional designation "TM-1," include: delayed production (2 to 10 days) of acid from glucose, formation of gas during nitrate reduction, and the frequent formation of "pits" in the agar surface. On TM, nonpitting colonies of TM-1 are morphologically similar to colonies of Neisseria gonorrhoeae and N. lactamica. Comparison of the characteristics of TM-1 strains with other similar fastidious gram-negative organisms encountered in clinical laboratories indicates that TM-1 is a distinct species. Further studies are required before proper taxonomic placement can be made.

GONORRHEA IN WOMEN

Two hundred thirty-one female patients of private obstetricians and gynecologists were examined for gonorrhea, using cultural techniques. Specimens were taken from the cervix and rectum of all patients and cultured on Thayer-Martin (TM) selective medium. Six, or 2.6%, were found positive. Of these, all were in the 20- to 29-year age group; only one had knowledge of being exposed to gonorrhea; three were pregnant. The frequently asymptomatic nature of gonorrhea in females, combined with a definable incidence of infection in this patient group, suggests that more frequent use of cultural methods employing TM medium would help control this rapidly increasing disease. Cultures are particularly recommended for women under 30 years who are single, pregnant, or manifest acute vaginitis.

Gonorrhea in Women

Two hundred thirty-one female patients of private obstetricians and gynecologists were examined for gonorrhea, using cultural techniques. Specimens were taken from the cervix and rectum of all patients and cultured on Thayer-Martin (TM) selective medium. Six, or 2.6%, were found positive. Of these, all were in the 20- to 29-year age group; only one had knowledge of being exposed to gonorrhea; three were pregnant. The frequently asymptomatic nature of gonorrhea in females, combined with a definable incidence of infection in this patient group, suggests that more frequent use of cultural methods employing TM medium would help control this rapidly increasing disease. Cultures are particularly recommended for women under 30 years who are single, pregnant, or manifest acute vaginitis.

Sensitivity and reproducibility of Thayer-Martin culture medium in diagnosing gonorrhea in women

The Thayer-Martin culture medium was evaluated by culturing specimens from the cervix and rectum of 59 women alleged to have been the source of gonorrhea in men. By testing these women on 3 consecutive days, 50 (85 per cent) were culturally confirmed to have gonorrhea. A single cervical culture detected 89.7 per cent of those infected; repeating a single cervical culture on a subsequent day resulted in a total detection of 96.6 per cent of those infected. The remaining 3.4 per cent in the detection rate was achieved by a single rectal culture. Reproducibility of positive results occurred in 91 per cent of 266 cervical specimens and 89 per cent of 165 rectal specimens. The Thayer-Martin culture medium provides an accurate and reliable means of detecting gonorrhea in women, and its use should be considered for routine screening of high-risk populations.