Clinical laboratory evaluation of automated microbial detection/identification system in analysis of clinical urine specimens.

More than 4,000 clinical urine specimens were evaluated with an automated microbial detection/identification system compared to a standarized manual analysis and the routine modalities used in five peer-group laboratories. The comparison indicates that the automated system recognizes the nine groups of significant microorganisms in urinary tract infections in hospitalized patients with the same efficiency as a standarized manual method. The automated system's ability to enumerate the bacterial populations in the original clinical specimen attained a high degree of accuracy.

Multi-laboratory evaluation of an automated microbial detection/identification system.

An automated and computerized system (Automicrobic System [AMS]) for the detection of frequently encountered bacteria in clinical urine specimens was tested in a collaborative study among six laboratories. The sensitivity, specificity, reliability, and reproducibility of the AMS were determined, and the system was compared with conventional detection and identification systems. In this study, pure cultures and mixtures of pure cultures were used to simulate clinical urine specimens. With pure cultures, the sensitivity of the AMS in identifying the nine groups of organisms most commonly found in urine averaged 92.8%. The specificity averaged 99.4%, and the reliability of a positive result averaged 92.1%. The latter value was strongly influenced by a relatively high occurrence of false positive Escherichia coli results. The AMS was capable of detecting growth of most organisms, including those which it was not designed to identify. However, it identified some of these incorrectly as common urinary tract flora. Reproducibility of results, both within laboratories and among different laboratories, was high. Fast-growing organisms, such as E. coli and Klebsiella/Enterobacter species, were detected often at cell populations well below the AMS enumeration threshold of 70,000/ml. In mixed culture studies, high levels of sensitivity and specificity were maintained but when Serratia species were present in mixtures with other organisms, there was often a false positive report of E. coli. The overall performance of the AMS was considered satisfactory under the test conditions used.

Unusual Enterobacteriaceae: H2S+ Shigella sonnei, one authentic and one false positive due to contamination with the obligate anaerobe Eubacterium lentum.

A mixture of Shigella sonnei and Eubacterium lentum produced H2S in triple sugar iron agar; however, neither produced any in pure culture. A second culture of S. sonnei, isolated in Japan, is thought to be the first documented H2S+ Shigella.

Effect of temperature on transport and plating media for enteric pathogens.

The effect of wide variations in incubation temperatures and long periods of incubation on transport and enrichment broths and plating media was determined by exhaustive analysis of 132 diarrheal stools for salmonellae and shigellae. Homogenized stools were streaked onto eosin methylene blue (EMB), Salmonella-Shigella (SS), and xylose lysine deoxycholate (XLD) agar plates, and into saline, Cary-Blair (CB) transport medium, and Selenite F and gram-negative (GN) enrichment broths. Incubation temperatures were compared at 20 C, 35 C, 40 C and ambient, and over a range of 4 to 52 C for media incubated in an insulated picnic cooler in an auto trunk. At 1, 2, 4, and 7 days the plates were observed, and the broths were subcultured. Each stool was streaked to 12 plates for 48 observations and pickings, and to 48 tubes, subcultured to 192 plates, for a total of 240 observations for pathogens. Analysis of data from 6,246 Salmonella-positive plates showed direct streaking to be most effective after 2 days of incubation, but broths were equally effective at 1 or 2 days. By day 4 many plates were overgrown, and both plates and broths showed diminution of positives by about 10% and at day 7, 19%. The 2,434 Shigella-positive plates were more demanding in all times and temperatures of incubation than salmonellae. Although at day 2 best results were obtained on direct streaking, shigellae die-offs in broths were excessive, with positive declining 23.7% by day 2, 49% by day 4, and 60% by day 7. Direct plating of both pathogens was poor at 20 C with about 48% success, but salmonellae preferred higher temperatures (35 and 40 C), whereas shigellae chose 35 C and ambient, which averaged 28 C for the 10-month study. Temperature was immaterial to salmonellae in broths with ambient slightly better than 35 C, but shigellae preferred 20 C and showed a 50% failure rate at 40 C, ambient being equal to 35 C. The preferential rank of broths in efficacy was GN greater than selenite greater than saline greater than CB greater than direct for salmonellae; for shigellae, GN greater than saline greater than direct greater than CB greater than selenite, with selenite proving to be unsuitable for shigellae. Plating media preferences were XLD greater than EMB greater than SS. Ten of 39 shigellae strains could not be recovered from the selenite and SS media combination, the many replications notwithstanding. The effectiveness of salmonellae and shigellae detection at ambient temperatures in Louisiana during the 10-month study period, as compared to controlled incubation temperatures, indicates that satisfactory enteric bacteriology can be done in warm climates without constant temperature incubators.

Effect of temperature of incubation on performance of media in the detection of enteric pathogens.

The effect of incubation temperatures on the efficacies of both plating media and transport or enrichment broths was determined by the analysis of 391 diarrheal stools for salmonellae and shigellae. Each analysis resulted in 90 observations. Stool specimens were homogenized in saline and used to inoculate eosin methylene blue (EMB), Salmonella-Shigella (SS), and xylose lysine deoxycholate (XLD) agar plates, Amies and Cary-Blair (CB) transport media, and gram-negative (GN) enrichment broth. All media were incubated at 25, 30, and 35 C for 24 and 48 h. In order of efficacy, GN and saline were significantly better than Amies and CB, which were still better than direct streaking for both salmonellae and shigellae. Forty-eight hours was a significant improvement over 24 h only at 25 C on direct streaking for both pathogens. Salmonella detection was also improved at 30 over 25 C on direct streaking. In direct plating, XLD was better than both SS and EMB for both pathogens. After broths, for salmonellae, XLD > SS > EMB, and for shigellae, XLD > EMB > SS, with all differences significant. SS agar was significantly improved for detection of shigellae with 48-h broth inocula versus 24-h broth inocula. The differences thus observed at the various temperatures tested proved to be less important than the media used. The efficient media, GN broth, saline-stool, and XLD were shown to be affected very little by either temperature or time variance of the magnitude tested.

Catalase test as an aid to the identification of Enterobacteriaceae.

Although the catalase test has been used for many years for rapid differentiation of the genera of gram-positive organisms, little has been said about its use in the family Enterobacteriaceae. It was further noted that a wide variety of methods exist for the execution of the catalase test, that there is no universally accepted strength specified for the hydrogen peroxide, and that no gradations for the vigor and speed of the reaction have been mentioned. Under the conditions of the clinical laboratory, we have developed a simple, rapid, and accurate method for the catalase test that has been of great value as an aid in the identification of the Enterobacteriaceae. With 3% H(2)O(2), it was observed that Serratia, Proteus, and Providencia were vigorous catalase reactors. Only Salmonella and rare Escherichia, Enterobacter, and Klebsiella isolates were moderate catalase reactors. Escherichia and Shigella strains were mostly nonreactive, with less than one-third weekly (+) reactive, whereas most Enterobacter strains tended to be weakly reactive. Klebsiella strains were divided equally between nonreactive and weakly reactive. In practice, this test was also of great value in discerning nonpigmented Serratia cultured from the hospital environment and in detecting mixed flora containing nonspreading Proteus.

Isolation of Shigellae. 8. Comparison of xylose lysine deoxycholate agar, hektoen enteric agar, Salmonella-Shigella agar, and eosin methylene blue agar with stool specimens.

Two enrichment broths and four plating media were compared for efficiency of detection of enteric pathogens from 1,597 stool specimens. Of 170 salmonellae isolated from the composite of all methods, direct streaking yielded but 54%, whereas enrichment in gram-negative broth found 87% and Selenite-F broth 97%. By contrast, gram-negative broth produced 100% of the 17 shigellae, Selenite-F broth but 77%, and direct streaking only 59%. Thus, enrichment methods produced almost twice the number of both pathogens as direct streaking. Comparison of the plating media revealed xylose lysine deoxycholate agar (XLD) and Hektoen enteric agar to be equal in their abilities to find both pathogens. Both were moderately better than Salmonella-Shigella agar and markedly superior to eosin methylene blue agar. XLD fround 83% of salmonellae produced by the composite of four media and 90% of the shigellae. Hektoen enteric agar found 80% of both. Salmonella-Shigella agar detected 74 and 68%, respectively, and eosin methylene blue agar only 42 and 63%. The numbers of false positives accruing to each medium, however, showed Hektoen enteric and Salmonella-Shigella agars to produce more than twice as many false-positive plates as XLD. Similarly, Selenite-F broth resulted in many more false-positives for all plating media than did gram-negative broth. Consequently, the index of validity, which equates successful isolation of pathogens with total pickings, favored XLD and gram-negative broth as the media of choice, with direct streaking the poorest method by all counts.

Group medical practice-an alternative to hospital oligopoly.

The aim of this article is to provoke dialogue and to initiate change. Costs of health care are alarming. Our inability to respond effectively to the demand for total health care is frustrating. Some drastic changes are required. This article, the first of a series that will enquire into our present methods for providing health care, suggests that in group medical practice some satisfactory solutions may be found.

The role of the general hospital in health care: part two in a series on group practice.

The aim of this article is to provoke dialogue and to initiate change. Costs of health care are alarming. Our inability to respond effectively to the demand for total health care is frustrating. This is the second article in a series on hospitals. The next article will look at unmet health care needs and underutilized health care resources and seek to identify the group practice of medicine as the logical centre for orientation of other than general hospital health services.

Isolation of Shigellae. VII. Comparison of gram-negative broth with Rappaport's enrichment broth.

Efficacies of Gram Negative Broth (GN) and Rappaport's Enrichment Broth (RE) were compared for detection of enteric pathogens from clinical specimens. Whereas direct streaking on four plating media found 57% of the salmonellae, GN found 80% and RE found 92% of the 157 isolates. By contrast, direct streaking found 87% of the shigellae whereas GN detected 93%, but RE found only 20%. RE produced 35% more Salmonella-positive plates than GN did, which resulted in a 15% increase in isolates. Statistically, RE proved to be significantly better than GN. The unsuitability of RE for shigellae, however, dictates that, if only one enrichment broth is to be used, GN must be that choice, but that maximal isolations of all enteric pathogens should result from the use of both RE and GN. Of the four plating media, Xylose Lysine Deoxycholate Agar detected 90% of the salmonellae and 85% of the shigellae. Salmonella-Shigella Agar detected 72 and 43%, respectively, and Levine Eosin Methylene Blue Agar found 45 and 55%. Bismuth Sulfite Agar detected only 22% of the salmonellae and found no shigellae. The performance of all of the plating media was enhanced by enrichment, but RE was especially effective for salmonellae when compared to GN.

Isolation of shigellae. V. Comparison of enrichment broths with stools.

Many enteric media are more efficient for the detection of salmonellae than of shigellae. Comparisons of three enrichment broths and three plating media were made during analysis of 1,405 stool specimens to choose a combination of media which would enhance detection of shigellae as well. Gram-Negative (GN), Selenite, and Silliker's Broths were streaked to E M B, Salmonella-Shigella (SS), and xylose lysine deoxycholate (XLD) Agars. The enrichment broths produced a twofold increase in isolations of both salmonellae and shigellae over direct streaking. All three broths performed equally well for Salmonella detection, but GN and Silliker's produced twice as many Shigella isolates as did Selenite. Comparison of the plating media showed that XLD was markedly more efficient than either E M B or SS Agar for the recovery of both genera. SS Agar was superior to E M B for isolation of salmonellae after enrichment, whereas E M B was better for isolation of shigellae by direct streaking. Both E M B and SS were more effective when used after GN and Silliker's than after Selenite. GN Broth and XLD Agar were the most efficient combination of media. During these analyses, 158 salmonellae and 49 shigellae isolates were obtained.

Isolation of shigellae. VI. Performance of media with stool speciments.

The efficiencies of three enrichment broths and four plating media for isolation of enteric pathogens were compared from 1,117 stool specimens. Direct streaking proved to be inferior to enrichment, detecting only 50% of the salmonellae and 61% of the shigellae. By contrast, Selenite Broth (SF) found 90% of the total salmonellae isolates and 82% of the shigellae isolates. Gram-Negative Broth (GN) found 82% and 85%, respectively, but Tetrathionate found only 60% and 39%. Thus, SF and GN were comparable for both salmonellae and shigellae and significantly better than Tetrathionate Broth for both. The plating media compared were MacConkey (MAC), deoxycholate citrate (DC), xylose lysine deoxycholate (XLD), and xylose lysine Brilliant Green (XLBG) Agars. Of the total salmonellae isolated, XLD produced 94%; XLBG, 71%; MAC, 55%; and DC, only 35%. Of shigellae, XLD found 89%; MAC, 75%; XLBG, 63%; and DC, but 27%. The efficacy of XLD is observed to be almost threefold that of DC. The most successful combination of media for the detection of fecal pathogens was GN or SF enrichment broths streaked to XLD plates. These analyses resulted in the isolation of 118 strains of salmonellae and 33 of shigellae.