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Biomedical subjects

M T Collins

Publications and source records attributed to M T Collins.

At least 91 records · Page 5Linked to original sources

An unusual strain of Legionella micdadei.

A microorganism antigenically identified as Legionella micdadei but showing a cellular fatty acid profile distinct from that described previously for this species and more similar to that of L. bozemanii has been studied by phenotypic characterization, crossed immunoelectrophoresis, gas-liquid chromatography, and transmission electron microscopy. Although the phenotypic characters, the crossed immunoelectrophoresis, and the ultrastructural features were similar to those of L. micdadei, the fatty acid composition differed significantly from this species; moreover it differed also from L. bozemanii, even though it was quantitatively more similar.

Antigens, Bacterial↗

Comparison of conventional and long-acting oxytetracyclines in prevention of induced Actinobacillus (Haemophilus) pleuropneumoniae infection of growing swine.

These experiments tested the hypothesis that long-acting oxytetracycline (oxytetracycline-LA) was more effective than regular oxytetracycline in preventing porcine pleuropneumonia when administered either 24 or 48 h prior to experimental challenge with virulent strains of Actinobacillus pleuropneumoniae. Two experiments (1 and 2) were conducted using growing pigs (average weight 12-15 kg). Antibiotic treatments were administered once intramuscularly at 20 mg/kg body weight; controls received an equivalent volume of saline. Clinical signs were recorded over seven days, and mortality rates and pathological lesions were analyzed using analysis of variance. Serum oxytetracycline levels were compared 48 and 72 h postinjection. All pigs developed clinical disease following experimental infection. Actinobacillus pleuropneumoniae was recovered from 42% of experiment 1 pigs and all of experiment 2 pigs. The data showed that both oxytetracycline and oxytetracycline-LA given at the same dose protected pigs against experimental infection when given 24 h prior to challenge, and there was no difference between the efficacy of the two drugs in this experiment. When administered 48 h prior to challenge, only oxytetracycline-LA reduced the clinical signs and pathological changes following A. pleuropneumoniae challenge. Between 48 and 72 h postinjection, oxytetracycline-LA blood levels were significantly greater compared to oxytetracycline-treated pigs.

Actinobacillus Infections↗

Cloning and expression of the Legionella micdadei "common antigen" in Escherichia coli.

To study individual Legionella antigens, a Legionella micdadei genomic library in Escherichia coli SC181 was established. Partially Sau3A digested L. micdadei DNA fragments (15-25 kilobase pairs (kb] were cloned into the tetracycline resistance gene of the cosmid vector pHC79. Four thousand ampicillin resistant recombinants were obtained; seven hundred were screened for expression of Legionella antigens in Western blot analysis with a polyspecific E. coli-absorbed anti-L. micdadei rabbit antibody. One of the positive clones expressed a 60 kilodalton (K) antigen, which reacted strongly with a monospecific rabbit antiserum raised against L. micdadei "common antigen" (CA), and an additional 13 K L. micdadei protein. The region encoding these two proteins from the 17 kb recombinant plasmid (pBA 2) was subcloned in pBGS18+. The DNA sequence of the CA encoding region in the 2.7 kb subcloned fragment will provide important information with respect to genetic vs. antigenic relatedness among Legionellae and other Gram-negative species, as well as to CA structure and possible function.

Antigens, Bacterial↗

Blood gas stability and hematological changes in experimentally-induced acute porcine pleuropneumonia.

Blood gas and hematological responses to acute, mild Actinobacillus pleuropneumoniae infection of growing pigs was studied. Six pigs (average weight 10.1 kg) were experimentally infected intranasally with A. pleuropneumoniae serotype 5. Four pigs served as controls. Rectal temperatures and arterial blood for gas analysis and hematology were taken at 0, 8, 16, 24, 48 and 72 h postinfection. All infected pigs became febrile showing clinical signs typical of mild to moderate porcine pleuropneumonia; controls remained asymptomatic. Neutrophilia with bands and lymphopenia were observed only in infected pigs. Arterial partial pressures of O2 and CO2, and pH did not change in infected pigs. All pigs were killed after 72 h, and lungs were examined and cultured. Gross and microscopic lesions consistent with porcine pleuropneumonia were seen in 3/6 and 5/6 infected lungs, respectively. Control lungs were grossly normal with no histological evidence of pleuropneumonia. We conclude that in mild, acute porcine pleuropneumonia as established experimentally, a leukogram typical of acute inflammation and stress is seen; however, hypoxemia and alveolar hypoventilation are not features of this form of the disease.

Actinobacillus Infections↗

Turbinate perimeter ratio as an indicator of conchal atrophy for diagnosis of atrophic rhinitis in pigs.

Transverse sections of snouts from 171 cross-bred (principally Yorkshire X American Landrace) pigs were evaluated for evidence of turbinate atrophy by use of conventional (atrophic rhinitis [AR] score) and morphometric methods. Of the 171 pigs, 35 were clinically normal (AR score, 0), 65 had mild AR (AR score, 1), 41 had moderate AR (AR score, 2), and 30 had severe AR (AR score, 3). Turbinate cross-sectional area (TA) and the ratio of TA to nostril cross-sectional area, called turbinate area ratio (TAR), had the lowest correlations (r = 0.24 to 0.55) with conventional AR score. Among clinically normal pigs, TA was greater in older pigs as expected, but the TAR values also were significantly (P less than 0.0001) different between 15-week-old pigs (55 kg) and 22-week-old pigs (100 kg). Turbinate perimeter and turbinate perimeter ratio (TPR) were not influenced by pig age or source. The TPR values were closely correlated with subjective visual AR scores (r = 0.73), with AR scores derived by measuring the space between the ventral portion of the scroll and the floor of the nasal cavity (r = 0.72), and the actual size of this space in millimeters (r = 0.71). Mean TPR values for pigs assigned visual AR scores of 0, 1, 2, or 3 were 1.54, 1.25, 0.97, and 0.73, respectively. The 95% confidence intervals around these mean TPR values were discreet and did not overlap. Turbinate perimeter ratio, therefore, may be a more reliable morphometric measure of atrophic rhinitis and also provides parametric data suitable for quantitative analysis.

Animals↗

A model for analyzing growth kinetics of a slowly growing Mycobacterium sp.

This report describes a simple method for quantifying viable mycobacteria and for determining generation time. We used statistical models and computer analysis of growth curves generated for the slowly growing mycobacterium Mycobacterium paratuberculosis under controlled conditions to derive a mathematical formula relating the dependent variable, growth, to the independent variables, log10 number of organisms in the inoculum (inoculum size) and incubation time. Growth was measured by a radiometric method which detects 14CO2 release during metabolism of a 14C-labeled substrate. The radiometric method allowed for early detection of growth and detected as few as three viable bacteria. The coefficient of variation between culture vials inoculated with the same number of M. paratuberculosis was 0.083. Radiometric measurements were highly correlated to spectrophotometric and plate count methods for measuring growth (r = 0.962 and 0.992, respectively). The proportion of the total variability explained by the model in a goodness of fit test was 0.9994. Application of the model to broth cultures provided accurate estimates of the number of M. paratuberculosis (standard error = 0.21, log10 scale) and the growth rate (coefficient of variation, 0.03). Generation time was observed to be dependent upon the number of organisms in the inoculum. The model accurately described all phases of growth of M. paratuberculosis and can likely be applied to other slowly growing microorganisms.

Carbon Radioisotopes↗

Antigenic analysis of Pasteurella haemolytica serovars 1 through 15 by crossed immunoelectrophoresis.

Pasteurella haemolytica serovars 1 through 12, grown in broth and on agar plates, and 2 field isolates (types A1 and T10) were used to develop polyvalent crossed immunoelectrophoresis (XIE) reference systems. The maximal number of antigens was revealed by XIE when sonicates of agar plate-grown organisms were used as the immunogen (to produce antibodies) and as the soluble antigen for XIE. Antigens produced from agar plate-grown organisms were less contaminated (by antigenic components of the medium) than were those produced from organisms grown in broth. Seventy-two antigens were detected in sonicated preparations of agar plate-grown P haemolytica. The common antigen of gram-negative bacteria was identified in the P haemolytica XIE reference system; precipitation was observed with rabbit antiserum to the common antigen of gram-negative bacteria isolated from Escherichia coli, as well as with rabbit immunoglobulins (obtained from unvaccinated rabbits). Most preimmune sera from our vaccinated rabbits also precipitated the common antigen. Serovar-specific antigens in the P haemolytica XIE reference system were defined and presumptively identified as part of the bacterial lipopolysaccharide complex by use of the limulus amebocyte lysate test. Partial cross-reactions were found between serovar-specific antigens within each biovar (A and T). Pasteurella haemolytica biovar A-specific and biovar T-specific antigens were defined by crossed-line immunoelectrophoresis. When serovars A13, A14, and T15 were tested in the P haemolytica XIE reference system, they gave high matching coefficient values of 0.98, 0.98, and 0.87, respectively. The proposal to separate P haemolytica biovars A and T into 2 different species was supported by immunotaxonomic data obtained from crossed immunoelectrophoresis, but more extensive studies will be necessary to establish the appropriate taxonomic position of these 2 groups of organisms.

Animals↗

Development of turbinate lesions and nasal colonization by Bordetella bronchiseptica and Pasteurella multocida during long-term exposure of healthy pigs to pigs affected by atrophic rhinitis.

Natural transmission of atrophic rhinitis from pigs from a herd with an endemic atrophic rhinitis problem to pigs from a herd free of atrophic rhinitis was demonstrated. Six replicates each with five pigs from the endemic atrophic rhinitis herd (Group A) and five pigs from the atrophic rhinitis-free herd (Group B) were housed together from 5 wk of age, with each replicate kept in isolation rooms maintained at optimal and controlled environmental conditions. Three replicates each with six pigs/room from the atrophic rhinitis-free herd (Group C), served as nonexposed controls. Group C pigs remained healthy and had no turbinate atrophy at either 10 or 17 wk of study (atrophic rhinitis score = 0 on a 0 to 3 scale). Group A pigs had a mean atrophic rhinitis score of 1.85 +/- 0.84, and group B pigs developed atrophic rhinitis to a mean score of 1.57 +/- 0.70. The isolation rate and quantity of Pasteurella multocida found on nasal swabs was directly related to lesions while those for Bordetella bronchiseptica were inversely related to turbinate atrophy. Of the various types of P. multocida evaluated, nontoxigenic type A and toxigenic type D were both directly related to atrophic rhinitis while nontoxigenic type D strains were not. No toxigenic type A P. multocida strains were isolated.

Animals↗

Characterization of Mycobacterium paratuberculosis by gas-liquid and thin-layer chromatography and rapid demonstration of mycobactin dependence using radiometric methods.

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 (12A; Johnston Laboratories, Inc., Towson, Md.) 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 degrees C and read daily with a BACTEC Model 301 (Johnston Laboratories). After 8 days of incubation, the growth index readings for the test broths containing mycobactin P were twice those of the 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.

Animals↗

Prevalence of Legionnaires' disease in pneumonia patients admitted to a Danish department of infectious diseases.

During a 14-month study period, 92 patients admitted to the University Clinic for Infectious Diseases with pneumonia were investigated to determine the prevalence and severity of Legionnaires' disease (LD). The diagnosis of LD was based on positive serology. Antibodies to 10 different legionella antigens--Legionella pneumophila serogroups 1-6, Fluoribacter (Legionella) bozemanae, F. dumoffii, F. gormanii, and Tatlockia (Legionella) micdadei--were measured by the microagglutination (MA) and indirect fluorescent antibody (IFA) techniques. LD was diagnosed in 22 patients showing a 4-fold or greater rise of MA titers. 10 patients showed a 4-fold or greater rise of IFA titer, 2 had standing high titer. One patient died. Legionella infection was the second most common cause of pneumonia. However, in half of the cases legionella infection occurred concomitantly with Streptococcus pneumoniae, Haemophilus influenzae, Mycoplasma pneumoniae, Chlamydia psittaci, or viral infections. All 22 LD cases were sporadic. LD had been contracted abroad by 6 patients. Two of the legionella pneumonias were hospital-acquired. Half of the LD patients were older than 60 years. The majority of cases occurred during the winter months. Neither clinical chemistry parameters nor clinical features could distinguish LD from other types of pneumonia.

Adolescent↗

Experimental reproduction of septicemic pasteurellosis in feedlot lambs: bacteriologic and pathologic examinations.

Septicemic pasteurellosis (SP) was induced in feedlot lambs. Twenty-eight lambs, randomly allotted into 7 groups, were given combinations of 3 treatments: (i) immunosuppression using hydrocortisone solubilized in dimethyl sulfoxide, (ii) rapid changes in feed, from 100% roughage to 90% concentrate, and (iii) oral inoculation of Pasteurella haemolytica biotype T. Feed changes and immunosuppression by hydrocortisone were needed for the production of SP. Pasteurella haemolytica inoculation was not necessary for induction of SP in all cases, indicating an endogenous source of infection. Clinical pathologic, bacteriologic, and gross and microscopic pathologic findings of induced SP were similar to those described for naturally occurring SP in lambs. Infection of lambs with P haemolytica biotype T via the gastrointestinal tract is discussed as a possible step in the pathogenesis of SP in feedlot lambs.

Animals↗

Effect of hydrocortisone on circulating lymphocyte numbers and their mitogen-induced blastogenesis in lambs.

The effect of hydrocortisone on the number of circulating lymphocytes and their blastogenic response was studied in 20 feedlot lambs given combinations of 3 treatments: hydrocortisone (25 mg/kg of body weight, 4 times a day, IM), feed changes (100% roughage to 90% concentrate over a 6-day period), and oral inoculation of Pasteurella haemolytica biotype T (10(9) to 10(11) bacteria/day via stomach tube) to develop a model for reproduction of septicemic pasteurellosis. Hydrocortisone caused lymphopenia and inhibited the blastogenic response of peripheral blood lymphocytes to phytohemagglutinin and concanavalin A mitogens. A synergistic effect was observed between hydrocortisone injections and feed changes resulting in higher than expected serum hydrocortisone concentrations and lower circulating lymphocyte counts. Seemingly, stress-induced increases in serum hydrocortisone concentrations cause suppression of the immune response of feedlot lambs. The combined effect of feed changes and stress on the immune response of lambs may explain the role of these 2 factors in the pathogenesis of septicemic pasteurellosis.

Animals↗

Niacin, nitrate and pyrazinamide studies using Middlebrook 7H10 Broth.

An innovative and rapid method for testing mycobacteria was developed using Middlebrook 7H10 Tween Broth in place of conventional media. Niacin production, nitrate reduction and the breakdown of pyrazinamidase were determined in 198 mycobacteria isolates. Less than nine days were required to obtain positive test results, and the correlation of tween broth with conventional test methods exceeded 98%.

Amidohydrolases↗

Comparison of a micro-agglutination test and the indirect immunofluorescence test for Legionella antibodies in patients.

A micro-agglutination (MA) test for antibodies to 10 different Legionella antigens was compared with the indirect immunofluorescence (IF) test. The antigens used were prepared from Legionella (L.) pneumophila serogroups (SG) 1-6, L. micdadei, L.bozemanii, L.dumoffii, and L.gormanii. Sera from 728 patients suspected of suffering from Legionella pneumonia were investigated by both tests. Thirty-five patients showed a significant rise in IF titre to greater than or equal to 128, and 43 had single or static titres of greater than or equal to 256 while the remaining 150 had negative IF titres. The results of the comparison indicate that the MA test offers at least the same sensitivity as the IF test in demonstrating diagnostic titre rises. It may be used as a screening test for selection of sera for further analysis by the IF test. In contrast to the IF test, MA is simple and inexpensive.

Agglutination Tests↗

Agglutinating antibody titers to members of the family Legionellaceae in cystic fibrosis patients as a result of cross-reacting antibodies to Pseudomonas aeruginosa.

The objective of this study was to evaluate the prevalence and significance of antibody titers to organisms in the family Legionellaceae in 128 serum samples collected from cystic fibrosis patients at routine examinations. Antibody titers were determined for 10 antigenic types of Legionellaceae; Legionella pneumophila serogroups 1 to 6, Fluoribacter (Legionella) bozemanae, Fluoribacter (Legionella) dumoffii, Fluoribacter (Legionella) gormanii, and Tatlockia (Legionella) micdadei. The method of antibody titer determination was the microagglutination test. Elevated titers (greater than or equal to 1:64) to one or more antigens were found in 41.3% of cystic fibrosis patients but in only 9.7% of 103 normal control subjects (P less than 0.01). Titers to 8 of the 10 antigens were directly correlated with the number of Pseudomonas aeruginosa precipitating antibodies in patient sera, as determined by crossed immunoelectrophoresis (correlation coefficients, greater than or equal to 0.74). Cross-reactions between P. aeruginosa and L. pneumophila were substantiated by crossed immunoelectrophoresis of hyperimmune rabbit serum as well as patient sera against P. aeruginosa and Legionellaceae antigens. Monospecific antibody to the "common antigen" of P. aeruginosa was used to demonstrate the presence of this antigen in L. pneumophila. The presence of cross-reacting antibodies in cystic fibrosis patients chronically infected with P. aeruginosa emphasizes the need for cautious interpretation of antibody titers to members of the family Legionellaceae.

Adolescent↗

Radiometric studies with gas-liquid and thin-layer chromatography for rapid demonstration of hemin dependence and characterization of Mycobacterium haemophilum.

Eight isolates of Mycobacterium haemophilum were evaluated by radiometric methods to determine whether this test system could support the growth of these organisms as well as demonstrate their growth requirements for iron complexes such as hemin, ferric ammonium citrate, and blood. In addition, gas-liquid and thin-layer chromatography were evaluated to determine whether these procedures could further differentiate M. haemophilum from other mycobacteria. During the initial 24 to 48 h, there was no significant difference between the radiometric test broths containing iron complexes and control broths without iron supplementation. After 48 h, the test growth index readings rapidly increased, and control broth readings leveled off and declined. The mean growth index reading of the test broths after 6 days of incubation was 100 times that of the controls. The mean incubation time with supplemented 7H10 agar was 17 days. The use of radiometric media resulted in the demonstration of hemin dependence by M. haemophilum significantly earlier than with 7H10 agar. Of the three supplements studied, whole blood provided the greatest growth rate, followed by ferric ammonium citrate and hemin. When 12 species of mycobacteria other than M. haemophilum were radiometrically evaluated, no isolate demonstrated an iron complex requirement. Gas-liquid and thin-layer chromatography procedures were able to rapidly differentiate M. haemophilum from the other 12 Mycobacterium species.

Carbon Dioxide↗

Serologic evidence of Legionella infection in horses.

The indirect fluorescent antibody test was used to examine 109 samples of equine sera randomly selected from serum pools. Results were compared with titers obtained by the microagglutination (MA) test. A high correlation (r = 0.89) was found between titers measured by the 2 tests. Blood samples were obtained serially from a total of 156 horses at a research farm and the sera were tested against Legionella pneumophila serogroups 1 through 4 using the MA test; 29 horses (19%) seroconverted to at least 1 serogroup of L pneumophila. The indirect fluorescent antibody test substantiated the results obtained by the MA test. Seroconversions in horses provided additional evidence that horses become naturally exposed to legionellae.

Agglutination Tests↗