Use of Hydrogenophaga pseudoflava penetration to quantitatively assess the impact of filtration parameters for 0.2-micrometer-pore-size filters.

Filters rated as having a 0.2-microm pore size (0.2-microm-rated filters) are used in laboratory and manufacturing settings for diverse applications of bacterial and particle removal from process fluids, analytical test articles, and gasses. Using Hydrogenophaga pseudoflava, a diminutive bacterium with an unusual geometry (i.e., it is very thin), we evaluated passage through 0.2-microm-rated filters and the impact of filtration process parameters and bacterial challenge density. We show that consistent H. pseudoflava passage occurs through 0.2-microm-rated filters. This is in contrast to an absence of significant passage of nutritionally challenged bacteria that are of similar size (i.e., hydrodynamic diameter) but dissimilar geometry.

Isolation and characterization of recombinant lambda gt11 bacteriophages expressing four different Mycobacterium intracellulare antigens.

Four bacteriophages expressing different immunoreactive recombinant Mycobacterium intracellulare antigens were isolated from a lambda gt11 library with monoclonal antibodies to M. intracellulare. These four antibodies reacted with native M. intracellulare proteins of 54, 43, 40/38, and 22 kilodaltons. Southern blot hybridizations with DNA probes prepared from insert fragments of these bacteriophages confirmed the M. intracellulare derivation of the inserts. The physical maps of the immunoreactive phages were deduced by restriction enzyme digestions. The molecular weights of the expressed recombinant antigens were determined by Western (immuno-) blotting.

Isolation and characterization of a recombinant lambda gt11 bacteriophage which expresses an immunoreactive Mycobacterium intracellulare protein in Escherichia coli.

Disseminated Mycobacterium avium-Mycobacterium intracellulare (M. avium complex) disease is a prevalent opportunistic infection in patients with acquired immune deficiency syndrome. Because of the increasing importance of this disease, an M. avium complex lambda gt11 expression library was prepared. We screened the library with an absorbed anti-M. intracellulare serum and identified a recombinant phage which expressed a 190-kilodalton beta-galactosidase-M. intracellulare fusion protein. Lysates containing the 190-kilodalton fusion protein evoked strong humoral and cell-mediated responses. The immunoreactivity of the M. intracellulare recombinant protein suggests that antigens isolated from the expression library may be useful as skin test, serodiagnostic, or immunoprophylactic reagents for M. avium complex disease.

Characterization of serologic and cell-mediated reactivity of a 38-kDa antigen isolated from Mycobacterium tuberculosis.

An antigen of Mycobacterium tuberculosis with an m.w. of 38,000 has been isolated by affinity chromatography using a monoclonal antibody. This antibody bound only to an antigen found in M. tuberculosis and Mycobacterium bovis BCG. The specificity of the antigen was tested in a vertical study by immunodetection on western blots reacted with hyperimmune sera against M. tuberculosis, M. bovis, and 10 other Mycobacterium species. The antigen was detected only by antisera to M. tuberculosis and M. bovis. Specificity in cell-mediated immunity was tested by skin tests in guinea pigs sensitized with M. tuberculosis, Mycobacterium intracellulare, and Mycobacterium kansasii and by lymphocyte proliferation tests. The 38-kDa antigen induced positive skin test reactions regardless of the Mycobacterium species used to sensitize the animal. The ability of the 38-kDa antigen to sensitize for cell-mediated immunity was tested by injecting mice with the 38-kDa antigen and challenging their lymphocytes in vitro with various mycobacterial antigens. Lymphocyte proliferation was observed in the presence of 38-kDa antigen, M. tuberculosis sonicate antigen, and tuberculin purified protein derivative and to M. kansasii and M. intracellulare. The 38-kDa antigen may contain a specific epitope detected by serology, but also contains epitopes that are cross-reactive for cellular immunity.

Salmonella regrowth in compost as influenced by substrate (salmonella regrowth in compost).

Composting can eliminate pathogenic organisms, including salmonellae, from sewage sludge. However, if salmonellae are present in the compost at undetectable levels or are inoculated into the compost by infected animals or from other sources, they may regrow presenting a health hazard for certain uses of compost. In this study, we examined dilute mineral-salt extracts of three composts from widely separate composting sites in the United States and found that they supported growth ofSalmonella typhimurium. From kinetic studies of the growth of the organism on these extracts, we concluded that each compost produced on extraction a single water-soluble substrate and that the substrates from the different composts were very similar, if not identical.

Prevalence of antibodies to Legionella species in a series of patients in Israel.

A collection of serum specimens from 77 patients at various hospitals or clinics in Israel was used to determine the usefulness of the enzyme-linked immunosorbent assay (ELISA) with a multivalent antigen for the detection of legionella antibodies. Rickettsial infection rather than legionellosis was suspected in most of these patients. The multivalent antigen was derived from Legionella pneumophila serogroups 1-6, L. bozemanii WIGA, and L. micdadei TATLOCK. A preliminary test of the multivalent antigen with specific rabbit antisera had shown that homologous reactions were not appreciably reduced in strength or specificity by the presence of the heterologous antigens. The results with the human sera revealed that 28 patients (36%) had reciprocal dilution titers greater than or equal to 1,280 and 43 (56%) had titers greater than or equal to 320. Tests with univalent antigens identified L. bozemanii as the only or principal antigen reacting with 13 of these sera. In contrast to the sera reacting with other legionella antigens, the great majority (11 of 13) of L. bozemanii-positive sera reacted also with Rickettsia typhi. The data suggest that most, but not all, reactions with L. bozemanii were elicited by a cross-reacting R. typhi antigen. These results were confirmed by cross-absorption tests.

Occurrence, growth, and suppression of salmonellae in composted sewage sludge.

Composted sewage sludge may be used to improve soil quality, but there remains some doubt concerning the microbiological safety of the product. Sewage sludge composts from 30 municipalities were sampled, and four samples (12%) contained salmonellae (two contained fewer than 0.3/g, and the other two had 21/g and 1.7 X 10(4)/g). All 30 composts were inoculated with salmonellae; the populations decreased at a specific death rate of about 0.15 h-1 over 24 h at 36 degrees C. In irradiation-sterilized composts inoculated with salmonellae, the salmonellae grew at a rate of 0.65 doublings per h for over 24 h. Growth and death rates were found to be moisture and flora associated. The growth or death rates for antibiotic-resistant salmonellae were not different from those of nonresistant strains. It was concluded that the active indigenous flora of compost establishes a homeostatic barrier to colonization by salmonellae, and in the absence of competing flora, reinoculated salmonellae may grow to potentially hazardous densities. The active microflora of moist composts eliminated contaminating salmonellae (10(5)/g) after 6 weeks.

Modified agar medium for detecting environmental salmonellae by the most-probable-number method.

Salmonellae in the environment remain a potential source of disease. Low numbers of salmonellae have been detected and enumerated from environmental samples by most-probable-number methods which require careful colony selection from a plated agar medium. A modified xylose lysine brilliant green medium was prepared to control the loss of selectivity caused by heating the brilliant green component. Added agar reduced colony spreading. The medium contained 47 g of xylose lysine agar base per liter; the agar content was adjusted to 2%, autoclaved, cooled to 50 degrees C, and then amended just before pouring to include H2S indicator and 7 ppm (7 ml of 1:1,000 brilliant green per liter) of unheated brilliant green dye. H2S-positive salmonellae were easily detected from sewage sludge compost to the exclusion of most other gram-negative bacteria. As a result, fewer non-salmonellae were picked for further most-probable-number analysis, greatly reducing the work load associated with the most-probable-number method. Direct plating was possible for enumerating salmonellae in laboratory composts containing ca. 10(3) or more salmonellae.

Bacteria associated with false-positive most-probable-number coliform test results for shellfish and estuaries.

Aerobic and facultatively anaerobic bacteria isolated from false-positive, presumptive, total coliform, most-probable-number tests of Chesapeake Bay oyster, water, and sediment samples were characterized and then classified by numerical taxonomy. A total of 538 bacterial strains clustered into 17 phena, the predominant groups of which were Enterobacteriaceae (including Escherichia coli), Aeromonas spp., and Bacillus spp. Bacillus spp. were recovered most frequently from sediment samples. Gas-producing strains which were not members of the Enterobacteriaceae were not isolated during this study. However, disproportionately large numbers of atypical and anaerogenic lactose-fermenting strains were encountered. We concluded that no single, specific bacterial group can be identified as being responsible for the false-positive reaction in the presumptive coliform test. Instead, the false-positive reaction is a result of complex interactions among various genera, representing predominantly bacteria other than coliforms.

Rate of occurrence of false-positive results from total coliform most-probable-number analysis of shellfish and estuaries.

The incidence of confirmed test, false-positive coliform most-probable-number results was compared with environmental parameters and was found to be inversely related to water temperature. It is concluded that the completed coliform test must be done when water temperatures drop below 15 degrees C.

An estuarine agar medium for enumeration of aerobic heterotrophic bacteria associated with water, sediment, and shellfish.

A plate count agar was formulated for use in bacteriological analysis of estuarine samples and was tested together with standard plate count agar and an estuarine salts yeast extract agar for growth of aerobic, heterotrophic bacteria in water, sediment, and oysters. The estuarine agar was found to be efficient for enumerating aerobic, heterotrophic bacterial populations of water, sediment, and oysters, and is recommended for plate counts of estuarine samples.

Aerobic and facultatively anaerobic bacteria associated with the gut of Canada geese (Branta canadensis) and whistling swans (Cygnus columbianus columbianus).

Aerobic and facultatively anaerobic bacteria from the intestinal tracts of swans and geese were isolated and characterized as part of a larger study of the microbiological effects of migratory waterfowl on water quality. A total of 356 isolates were identified by using rapid identification methods and classified by using numerical taxonomy. A diverse population of bacteria was recovered from the waterfowl, and representative strains could be classified into 21 phena. The majority of the aerobic, heterotrophic bacteria found in the gut of the waterfowl were species of Enterobacteriaceae. Streptococcus. Lactobacillus, and Bacillus. Unfortunately, the birds that were examined did not harbor significant numbers of any waterfowl-specific bacterial species. Thus, it may not be possible to assess microbiological impact of migratory waterfowl by using and "indicator" species since avian fecal pollution could not be distinguished from animal and human fecal pollution.

Microbial impact of Canada geese (Branta canadensis) and whistling swans (Cygnus columbianus columbianus) on aquatic ecosystems.

Quantitative and qualitative analyses of the intestinal bacterial flora of Canada geese and whistling swans were carried out with the finding that wild birds harbor significantly more fecal coliforms than fecal streptococci. The reverse was typical of captive and fasting birds. Neither Salmonella spp. nor Shigella spp. were isolated from 44 migratory waterfowl that were wintering in the Chesapeake Bay region. Enteropathogenic Escherichia coli were detected in seven birds. Geese eliminated 10(7) and swans 10(9) fecal coliforms per day. Results of in situ studies showed that large flocks of waterfowl can cause elevated fecal coliform densities in the water column. From the data obtained in this study, it is possible to predict the microbial impact of migratory waterfowl upon aquatic roosting sites.