skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Enhanced radiometric detection of Mycobacterium paratuberculosis by using filter-concentrated bovine fecal specimens

Abstract

A commercial radiometric medium, BACTEC 12B, was modified by addition of mycobactin, egg yolk suspension, and antibiotics (vancomycin, amphotericin B, and nalidixic acid). Decontaminated bovine fecal specimens were filter concentrated by using 3-microns-pore-size, 13-mm-diameter polycarbonate filters, and the entire filter was placed into the radiometric broth. Comparison of the radiometric technique with conventional methods on 603 cattle from 9 Mycobacterium paratuberculosis-infected herds found that of 75 positive specimens, the radiometric technique detected 92% while conventional methods detected 60% (P less than 0.0005). Only 3.9% of radiometric cultures were contaminated. To measure the effect of filter concentration of specimens on the detection rate, 5 cattle with minimal and 5 with moderate ileum histopathology were sampled weekly for 3 weeks. M. paratuberculosis was detected in 33.3% of nonfiltered specimens and 76.7% of filtered specimens (P less than 0.005). Detection rates were directly correlated with the severity of disease, and the advantage of specimen concentration was greatest on fecal specimens from cattle with low-grade infections. Detection times were also correlated with infection severity: 13.4 +/- 5.9 days with smear-positive specimens, 27.9 +/- 8.7 days with feces from cows with typical subclinical infections, and 38.7 +/- 3.8 days with fecal specimens from cows withmore » low-grade infections. Use of a cocktail of vancomycin, amphotericin B, and nalidixic acid for selective suppression of nonmycobacterial contaminants was better than the commercial product PANTA (Becton Dickinson Microbiologic Systems, Towson, Md.) only when specimens contained very low numbers of M. paratuberculosis.« less

Authors:
; ; ; ; ;  [1]
  1. (Univ. of Wisconsin, Madison (USA))
Publication Date:
OSTI Identifier:
6070130
Resource Type:
Journal Article
Resource Relation:
Journal Name: Journal of Clinical Microbiology; (USA); Journal Volume: 28:11
Country of Publication:
United States
Language:
English
Subject:
62 RADIOLOGY AND NUCLEAR MEDICINE; FECES; RADIOMETRIC ANALYSIS; TUBERCULOSIS; DIAGNOSIS; CATTLE; CULTURE MEDIA; FILTRATION; MYCOBACTERIUM TUBERCULOSIS; SENSITIVITY ANALYSIS; ANIMALS; BACTERIA; BACTERIAL DISEASES; BIOLOGICAL MATERIALS; BIOLOGICAL WASTES; CHEMICAL ANALYSIS; DISEASES; DOMESTIC ANIMALS; INFECTIOUS DISEASES; MAMMALS; MATERIALS; MICROORGANISMS; MYCOBACTERIUM; QUANTITATIVE CHEMICAL ANALYSIS; RUMINANTS; SEPARATION PROCESSES; VERTEBRATES; WASTES; 550601* - Medicine- Unsealed Radionuclides in Diagnostics

Citation Formats

Collins, M.T., Kenefick, K.B., Sockett, D.C., Lambrecht, R.S., McDonald, J., and Jorgensen, J.B.. Enhanced radiometric detection of Mycobacterium paratuberculosis by using filter-concentrated bovine fecal specimens. United States: N. p., 1990. Web.
Collins, M.T., Kenefick, K.B., Sockett, D.C., Lambrecht, R.S., McDonald, J., & Jorgensen, J.B.. Enhanced radiometric detection of Mycobacterium paratuberculosis by using filter-concentrated bovine fecal specimens. United States.
Collins, M.T., Kenefick, K.B., Sockett, D.C., Lambrecht, R.S., McDonald, J., and Jorgensen, J.B.. 1990. "Enhanced radiometric detection of Mycobacterium paratuberculosis by using filter-concentrated bovine fecal specimens". United States. doi:.
@article{osti_6070130,
title = {Enhanced radiometric detection of Mycobacterium paratuberculosis by using filter-concentrated bovine fecal specimens},
author = {Collins, M.T. and Kenefick, K.B. and Sockett, D.C. and Lambrecht, R.S. and McDonald, J. and Jorgensen, J.B.},
abstractNote = {A commercial radiometric medium, BACTEC 12B, was modified by addition of mycobactin, egg yolk suspension, and antibiotics (vancomycin, amphotericin B, and nalidixic acid). Decontaminated bovine fecal specimens were filter concentrated by using 3-microns-pore-size, 13-mm-diameter polycarbonate filters, and the entire filter was placed into the radiometric broth. Comparison of the radiometric technique with conventional methods on 603 cattle from 9 Mycobacterium paratuberculosis-infected herds found that of 75 positive specimens, the radiometric technique detected 92% while conventional methods detected 60% (P less than 0.0005). Only 3.9% of radiometric cultures were contaminated. To measure the effect of filter concentration of specimens on the detection rate, 5 cattle with minimal and 5 with moderate ileum histopathology were sampled weekly for 3 weeks. M. paratuberculosis was detected in 33.3% of nonfiltered specimens and 76.7% of filtered specimens (P less than 0.005). Detection rates were directly correlated with the severity of disease, and the advantage of specimen concentration was greatest on fecal specimens from cattle with low-grade infections. Detection times were also correlated with infection severity: 13.4 +/- 5.9 days with smear-positive specimens, 27.9 +/- 8.7 days with feces from cows with typical subclinical infections, and 38.7 +/- 3.8 days with fecal specimens from cows with low-grade infections. Use of a cocktail of vancomycin, amphotericin B, and nalidixic acid for selective suppression of nonmycobacterial contaminants was better than the commercial product PANTA (Becton Dickinson Microbiologic Systems, Towson, Md.) only when specimens contained very low numbers of M. paratuberculosis.},
doi = {},
journal = {Journal of Clinical Microbiology; (USA)},
number = ,
volume = 28:11,
place = {United States},
year = 1990,
month =
}
  • Thirty-six Mycobacterium paratuberculosis isolates of bovine, caprine, and ovine origins were evaluated by using gas-liquid chromatography (GLC), thin-layer chromatography (TLC), and BACTEC 7H12 Middlebrook TB medium in an effort to more rapidly differentiate this group of organisms from other mycobacteria. Bacterial suspensions (0.1 ml) were inoculated by syringe into 7H12 broth containing 2 micrograms of mycobactin P per ml and control broth without mycobactin P. Cultures were incubated at 37/sup 0/C and read daily with a BACTEC Model 301. After 8 days of incubation, the growth index readings for the test broths containing mycobactin P were twice those of themore » control broths without mycobactin P. Sixty-five isolates of mycobacteria other than M. paratuberculosis were also examined. No difference was noted between the growth index readings of control and mycobactin-containing broths. Except for Mycobacterium avium-Mycobacterium intracellulare, TLC studies differentiated M. paratuberculosis from the other mycobacterial species tested. The GLC data reveal that all M. paratuberculosis isolates had a distinctive peak (14A) which was not found among M. avium-M. intracellulare complex organisms. These data indicate that 7H12 radiometric broth was able to rapidly demonstrate the mycobactin dependence of M. paratuberculosis and GLC and TLC procedures were capable of rapidly differentiating this organism from the other mycobacteria studied.« less
  • The formulation of media for selective, automatable, radiometric detection of growth of Mycobacterium tuberculosis in vitro is described. Palmitic-1 acid labeled with carbon-14 and formic-/sup 14/C acid were compared as substrate sources of (/sup 14/C)O2 in media deficient in carbohydrate and containing appropriate antimicrobial agents that are not active against tubercle bacilli. A preliminary clinical laboratory study of a medium containing 4 microCi palmitic-1-/sup 14/C acid per ml showed that this method might provide the basis for practical laboratory use.
  • A reconsideration of the laboratory methods used for primary isolation of mycobacteria other than Mycobacterium tuberculosis is needed due to the increasingly recognized importance of such mycobacterial infections in immunocompromised patients. One example of this is the severe opportunistic infections caused by Mycobacterium avium complex among AIDS patients. In this study, the Bactec radiometric system was compared to conventional culture on solid medium for the detection of M. avium complex in 3,612 selected clinical specimens, mainly of extrapulmonary origin. Of a total number of 63 M. avium complex isolates, the Bactec system detected 58 (92%), compared to 37 (59%) formore » conventional culture. A much more rapid detection was attained with radiometric technique than with conventional culture. The mean detection time for the cultures positive with both methods was 7.1 and 28.3 days, respectively. The Bactec radiometric system achieves a rapid and significantly more sensitive detection and seems to be an excellent complement to conventional culture in the laboratory diagnosis of infections with the M. avium complex.« less
  • Interest in rapid bacterial detection methods for sanitary indicator bacteria in water prompted a study of the use of (U-14C)mannitol to detect fecal coliforms (FC). A simple method which used m-FC broth, membrane filtration, and two-temperature incubation (35 degrees C for 2 h followed by 44.5 degrees C for 2.5 h) was developed. (U-14C)mannitol was added to the medium, and the temperature was raised to 44.5 degrees C after 2 h at 35 degrees C. 14CO2 was collected as Ba14CO3 and assayed by liquid scintillation spectroscopy. Correlations were examined between FC cell numbers at the start of incubation (standard 24-hmore » FC test) and Ba14CO3 counts per minute after 4.5 h. Results indicated that FC numbers ranging from 1 x 10(1) to 2.1 x 10(5) cells could be detected in 4.5 h. Within-sample reproducibility at all cell concentrations was good, but sample-to-sample reproducibility was variable. Comparisons between m-FC broth and m-FC broth modified by substituting D-mannitol for lactose indicated that the standard m-FC broth was the better test medium. Results from experiments in which dimethyl sulfoxide was used to increase permeability of FC to (U-14C)mannitol indicated no increase in 14CO2 production due to dimethyl sulfoxide. Detection of FC by this method may be useful for rapid estimation of FC levels in freshwater recreational areas, for estimating the quality of potable source water, and potentially for emergency testing of potable water, suspected of contamination due to distribution line breaks or cross-connections.« less