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The variable response of bacteria to excess ferric iron in host tissues.

The enhancement by exogenous ferric iron, both systemic and local, of the infectivity of 120 strains of bacteria, representing 17 genera, was measured in the skin of guinea-pigs. Systemic iron enhanced only 23% of 115 strains, and local iron 49% of 71 strains. Systemic iron, by an apparently anti-inflammatory action, depressed the size of lesions produced by 27 of the non-enhanced strains from nine of the genera tested. For most strains, the degree of enhancement was small, ranging from 2- to 8-fold, and often evident only with the more effective local iron; among these were some near-saprophytes like Mycobacterium phlei, M. smegmatis, Bacillus cereus and Clostridium bifermentans. Substantial enhancement, from 14- to 50-fold, was observed with the more pathogenic among the strains tested: namely BCG, Corynebacterium ovis, C. murium, Listeria monocytogenes, Erysipelothrix rhusiopathiae, Cl. perfringens, Cl. septicum, Cl. oedematiens, and some strains of Klebsiella spp., Proteus spp. and Aeromonas hydrophila. The enhancement of BCG by a single dose of iron given locally with the inoculum was only feebly manifest after 7 days, but substantial after 14--19 days, indicating the decisive effect of interference with an early humoral defence on the establishment of chronic infection some time later. Insofar as guinea-pigs whose antibacterial defences are lowered by substantial amounts of exogenous iron in the circulation represent human subjects at risk of infection because of clinical states characterised by excess of available iron, the results of the survey suggest that only a minority among the environmental bacteria can take advantage of the decreased resistance associated with such states; but that this minority is likely to include the more virulent strains in the environment.

Animals↗

The antagonism of tetracycline and ferric iron in vivo.

To test the hypothesis that the in-vivo antibiotic action of tetracycline might be affected by ferric iron and the enhancement of infection by ferric iron by tetracycline, the actions of intraperitoneal antibiotic and local ferric ammonium citrate, given separately and together, were measured in the dorsal skin of guinea-pigs bearing lesions due to staphylococci, streptococci, a Proteus sp., an Erysipelothrix sp., Clostridium perfringens, Pseudomonas aeruginosa, Aeromonas hydrophila and Klebsiella pneumoniae. Tetracycline, given in two intraperitoneal doses of 25 mg/kg at 0 and 2 h after intracutaneous challenge, maintained plasma concentrations of 4-6 micrograms/ml for more than the first 4 h of infection, after which the local lesions had become largely insusceptible to the antibiotic. The intracutaneous injection of Fe 10 micrograms in a volume of 0.1 ml containing the bacteria was sufficient to enhance infection by those strains susceptible to this effect. The in-vivo efficacy of tetracycline was not always related to low MIC; a low MIC was sometimes associated with little action and a high MIC with moderate action. Sixteen organisms were tested. The iron diminished the tetracycline effect only feebly with one staphylococcal strain and the strain of E. rhusiopathiae. In only one case, with a strain of Proteus sp., was the tetracycline action grossly diminished. On the other hand, tetracycline diminished the enhancement effect of iron moderately with three strains of staphylococci and one strain each of K. pneumoniae, P. aeruginosa and C. perfringens, and strongly with two strains of staphylococci, a group-C streptococcus and one strain each of K. pneumoniae, E. rhusiopathiae and A. hydrophila. It is evident that the diminution of tetracycline action by moderate excess of readily available Fe , whether endogenous or administered, is an unlikely event (three instances among the 16 tested) whereas the diminution of the infection-enhancing effect of iron by tetracycline is much more likely (12 instances among the 16). Insofar as a decrease in iron available for enhancement of infection is valid evidence of a diminution of the iron available for necessary physiological processes of the subject treated, our results suggest that these processes might be affected by tetracycline.

Animals↗

Phylogenic and phenotypic characterization of some Eubacterium-like isolates from human feces: description of Solobacterium moorei Gen. Nov., Sp. Nov.

Three isolated strains from human feces were characterized by biochemical tests and 16S rDNA analysis. Phylogenetic analysis revealed that these isolated strains were members of the Clostridium subphylum of gram-positive bacteria. The phenotypic characters resembled those of the genus Eubacterium, but these strains were shown to be phylogenetically distant from the type species of the genus, Eubacterium limosum. The strains showed a specific phylogenetic association with Holdemania filiformis and Erysipelothrix rhusiopathiae. Based on a 16S rDNA sequence divergence of greater than 12% with H. filiformis and E. rhusiopathiae, a new genus, Solobacterium, is proposed for three strains, with one species, Solobacterium moorei. The type strain of Solobacterium moorei is JCM 10645T.

Bacteria, Anaerobic↗

Microbiological status of piggery effluent from 13 piggeries in the south east Queensland region of Australia.

AIMS: To assist in the development of safe piggery effluent re-use guidelines by determining the level of selected pathogens and indicator organisms in the effluent ponds of 13 south-east Queensland piggeries. METHODS AND RESULTS: The numbers of thermotolerant coliforms, Campylobacter jejuni/coli, Erysipelothrix rhusiopathiae, Escherichia coli, Salmonella and rotavirus were determined in 29 samples derived from the 13 piggeries. The study demonstrated that the 13 final effluent ponds contained an average of 1.2 x 10(5) colony-forming units (CFU) 100 ml(-1) of thermotolerant coliforms and 1.03 x 10(5) CFU 100 ml(-1) of E. coli. The Campylobacter level varied from none detectable (two of 13 piggeries) to a maximum of 930 most probable number (MPN) 100 ml(-1) (two of 13 piggeries). Salmonella was detected in the final ponds of only four of the 13 piggeries and then only at a low level (highest level being 51 MPN 100 ml(-1)). No rotavirus and no Erysip. rhusiopathiae were detected. The average log10 reductions across the ponding systems to the final irrigation pond were 1.77 for thermotolerant coliforms, 1.71 for E. coli and 1.04 for Campylobacter. CONCLUSIONS: This study has provided a baseline knowledge on the levels of indicator organisms and selected pathogens in piggery effluent. SIGNIFICANCE AND IMPACT OF THE STUDY: The knowledge gained in this study will assist in the development of guidelines to ensure the safe and sustainable re-use of piggery effluent.

Animal Husbandry↗

Parvovirus antibodies in vaccinated gilts in field conditions--results with HI and ELISA tests.

This study was conducted to determine the antibody response for porcine parvovirus (PPV) of 39 gilts in field conditions after vaccination. Gilts from four herds endemically infected with PPV were injected twice with a commercial vaccine of inactivated PPV and Erysipelothrix rhusiopathiae. The PPV antibodies were analysed both with haemagglutination inhibition (HI) and enzyme-linked immunosorbent assay (ELISA) in order to study the agreement between these methods. The possible association between high-antibody titres and reproductive failure (repeat breeding, culling for infertility, < or = 6 piglets born alive) was also investigated. In these study herds, endemically infected by PPV, most gilts (84.6%) had not seroconverted by the age of 6 months. On-field vaccination resulted in a consistent increase of humoral immunity not exceeding the antibody level of 1 : 512 in the majority of gilts in all herds examined. The agreement between ELISA and HI tests was moderate (Spearman's rho = 0.87, kappa = 0.63). The seroconversion over the level >1:512 by mid-pregnancy was not associated with reproductive failure.

Agglutination Tests↗

Enterococcal-type glycopeptide resistance genes in non-enterococcal organisms.

Although the emergence of vancomycin-resistant enterococci can be attributed, in part, to the increasing use of vancomycin in clinical practice, and glycopeptide use in animal husbandry, the origins of the enterococcal vancomycin resistance genes are not clear. The vancomycin resistance-associated genes in Enterococcus gallinarum, Enterococcus casseliflavus/flavescens, Lactobacillus spp., Leuconostoc spp., Pediococcus spp., and Erysipelothrix rhusiopathiae, are not the source of the high-level vancomycin resistance-associated genes in enterococci. There are, however, environmental organisms which have been found to have gene clusters homologous to the enterococcal vanA, vanB and vanC gene clusters; these include the biopesticide Paenibacillus popilliae, and, to a lesser extent, the glycopeptide-producing organisms Amycolatopsis orientalis and Streptomyces toyocaensis. Still, the exact sources of the enterococcal vancomycin resistance genes remain a mystery.

Anti-Bacterial Agents↗

Grouping of some clinically relevant gram-positive rods by automated fatty acid analysis. Diagnostic implications.

One hundred and ninety-four strains of aerobically growing Gram-positive rods of the genera Corynebacterium, Actinomyces, Arcanobacterium, Erysipelothrix, Listeria, Oerskovia, Nocardia, Rhodococcus, and of unnamed Center for Disease Control (CDC) groups were checked for cellular fatty acid profiles with the microbial Identification System (Microbial ID, Newark, Del., USA). In order to obtain unified data usable for the clinical laboratory, 24 or 48 h sheep blood agar cultures were used. It was thought that grouping and perhaps identification could be aided by this approach. With the aid of numerical analysis, four groups (two consisting of two subgroups each) were established. The discriminatory ability of the scheme, however, was only 77.2%, indicating that grouping of an unknown isolate could be done with some accuracy, but that speciation would still require biochemical testing.

Bacterial Infections↗

In vitro susceptibilities of aerobic and facultative non-spore-forming gram-positive bacilli to HMR 3647 (RU 66647) and 14 other antimicrobials.

The comparative in vitro activity of the ketolide HMR 3647 (RU 66647) and those of structurally related macrolide-lincosamide-streptogramin compounds (erythromycin, roxithromycin, azithromycin, clarithromycin, josamycin, lincomycin, pristinamycin, and quinupristin-dalfopristin) as well as those of benzylpenicillin, doxycycline, vancomycin, teicoplanin, levofloxacin, and rifapentine against 247 aerobic and facultative non-spore-forming gram-positive bacilli were determined by an agar dilution method. The ketolide was active against most organisms tested except Corynebacterium striatum, coryneform CDC group 12, and Oerskovia spp. The frequency of resistance to erythromycin and other macrolides as well as that to lincomycin was high. Pristinamycin and, to a lesser extent, quinupristin-dalfopristin were very active, but resistance to these agents was present in some strains of Rhodococcus equi, Listeria spp., C. striatum, Erysipelothrix rhusiopathiae, and Oerskovia spp. HMR 3647 was very active against all erythromycin-sensitive and many erythromycin-nonsusceptible strains, especially Corynebacterium minutissimum, Corynebacterium pseudodiphtheriticum, Corynebacterium amycolatum, and Corynebacterium jeikeium. In vitro resistance to benzylpenicillin was common, but doxycycline, vancomycin, and teicoplanin were very active against most organisms tested except E. rhusiopathiae, against which glycopeptide antibiotics were not active. The in vitro activity of levofloxacin was remarkable, but resistance to this agent was common for C. amycolatum, Corynebacterium urealyticum, C. jeikeium, and Oerskovia spp. strains. Rifapentine was also very active in vitro against many organisms, but resistance to this agent was always present in E. rhusiopathiae and was very common in C. striatum and C. urealyticum.

Anti-Bacterial Agents↗

Rapid confirmation of Listeria monocytogenes isolated from foods by a colony blot assay using a digoxigenin-labeled synthetic oligonucleotide probe.

An oligodeoxyribonucleotide probe based on the sequence of a 321-bp internal fragment of the msp gene encoding a major secreted polypeptide of Listeria monocytogenes was labeled with digoxigenin by using terminal deoxynucleotidyl transferase. The specificity of the digoxigenin-labeled probe was determined by dot blot assays. The probe reacted with all strains of L. monocytogenes tested (12 of 12 strains representing five serotypes). The probe did not react with any other Listeria species or with other gram-positive bacteria (Brochothrix, Erysipelothrix, Corynebacterium, Rhodococcus, Lactobacillus, Leuconostoc, Bacillus, Staphylococcus, and Streptococcus). The probe was used to develop a colony blot assay for the rapid confirmation of L. monocytogenes on Listeria-selective agars which had been streaked with food enrichment cultures. Forty-eight food samples were tested by conventional culture and DNA colony blot assay. The sensitivity and specificity of the DNA colony blot were 100 and 97%, respectively.

Bacterial Proteins↗

Characteristics of a bacteriocin derived from Streptococcus faecalis var. zymogenes antagonistic to Diplococcus peumoniae.

A bacteriocin-producing strain of Streptococcus faecalis var. zymogenes (E-1) was isolated from clinical material (conjunctiva). The active substance differed from bacteriocins described by other investigators primarily in its spectrum of antibacterial activity, especially by its marked inhibition of Diplococcus pneumoniae. The E-1 bacteriocin also inhibited nonhemolytic strains of enterococci as well as one-third of the Viridans group of streptococcal strains investigated. The degree of inhibition, however, as indicated by the size of the zones against the latter organisms, was significantly reduced. No activity was detected against any of the strains belonging to the following groups of bacteria: hemolytic enterococci, beta-hemolytic streptococci, nonhemolytic streptococci, staphylococci, and various gram-negative species. Similarly, three strains each of Bacillus cereus and Listeria monocytogenes and one strain of Erysipelothrix insidiosa were not inhibited. The bacteriocin was able to diffuse through bacterial membranes as well as cellulose dialyzer tubing. It was inactivated by heating to 80 C for 20 min but resisted inactivation by either trypsin or chloroform.

Agar↗

Endocarditis due to rare and fastidious bacteria.

The etiologic diagnosis of infective endocarditis is easily made in the presence of continuous bacteremia with gram-positive cocci. However, the blood culture may contain a bacterium rarely associated with endocarditis, such as Lactobacillus spp., Klebsiella spp., or nontoxigenic Corynebacterium, Salmonella, Gemella, Campylobacter, Aeromonas, Yersinia, Nocardia, Pasteurella, Listeria, or Erysipelothrix spp., that requires further investigation to establish the relationship with endocarditis, or the blood culture may be uninformative despite a supportive clinical evaluation. In the latter case, the etiologic agents are either fastidious extracellular or intracellular bacteria. Fastidious extracellular bacteria such as Abiotrophia, HACEK group bacteria, Clostridium, Brucella, Legionella, Mycobacterium, and Bartonella spp. need supplemented media, prolonged incubation time, and special culture conditions. Intracellular bacteria such as Coxiella burnetii cannot be isolated routinely. The two most prevalent etiologic agents of culture-negative endocarditis are C. burnetti and Bartonella spp. Their diagnosis is usually carried out serologically. A systemic pathologic examination of excised heart valves including periodic acid-Schiff (PAS) staining and molecular methods has allowed the identification of Whipple's bacillus endocarditis. Pathologic examination of the valve using special staining, such as Warthin-Starry, Gimenez, and PAS, and broad-spectrum PCR should be performed systematically when no etiologic diagnosis is evident through routine laboratory evaluation.

Animals↗

EFFECTS OF CARBON DIOXIDE ON THE COLONIAL GROWTH OF PATHOGENIC LEPTOSPIRAE.

Yanagawa, R. (National Institute of Animal Health, Kodaira, Tokyo, Japan), T. Hiramune, and J. Fujita. Effects of carbon dioxide on the colonial growth of pathogenic leptospirae. J. Bacteriol. 85:875-880. 1963.-The growth of colonies of pathogenic leptospirae was examined in air of various CO(2) concentrations, ranging from air from which CO(2) was absorbed to air with 20% CO(2) added, and was compared with the growth of other microorganisms of the genera Mycoplasma, Brucella, Vibrio, Erysipelothrix, Staphylococcus, Micrococcus, Sarcina, Klebsiella, Salmonella, Escherichia, and Bacillus. The colonial growth of the 24 strains of pathogenic leptospirae examined was similar to that of other organisms in that the growth was negative in air from which naturally contained CO(2) had been absorbed, the growth tended to be inhibited in 5 to 20% CO(2) in proportion to the CO(2) concentration, and good colonial growth of leptospirae was obtained in a relatively narrow range of CO(2) concentration (from the normal amount in air to around 1%). The growth of some leptospirae was accelerated in 1% CO(2), but not that of the strains of Leptospira icterohaemorrhagiae, which required a lower range of CO(2), and of L. canicola. The CO(2) requirement of pathogenic leptospirae was similar, to some extent, to that of Mycoplasma mycoides, but was not so high as that of Brucella abortus. Incorporation of C(14)O(2) was demonstrated in leptospirae.

Animals↗

Cellular fatty acid composition as an adjunct to the identification of asporogenous, aerobic gram-positive rods.

Cellular fatty acid (CFA) compositions of 561 asporogenous, aerobic gram-positive rods were analyzed by gas-liquid chromatography as an adjunct to their identification when grown on blood agar at 35 degrees C. The organisms could be divided into two groups. In the first group (branched-chain type), which included coryneform CDC groups A-3, A-4, and A-5; some strains of B-1 and B-3; "Corynebacterium aquaticum"; Brevibacterium liquefaciens; Rothia dentocariosa; and Listeria spp., the rods had sizable quantities of antiesopentadecanoic (Ca15:0) and anteisoheptadecanoic (Ca17:0) acids. Other species with these types of CFA included B. acetylicum, which contained large amounts of isotridecanoic (Ci13:0) and anteisotridecanoic (Ca13:0) acids. CFAs useful for distinguishing among Jonesia denitrificans, Oerskovia spp., some strains of CDC groups B-1 and B-3, Kurthia spp., and Propionibacterium avidum were hexadecanoic (C 16:0) acid, isopentadecanoic (Ci15:0) acid, and Ca15:0). The second group (straight-chained type), which included Actinomyces pyogenes; Arcanobacterium haemolyticum; C. bovis; C. cystitidis; C. diphtheriae; C. flavescens, "C. gentalium"; C. jeikeium; C. kutscheri; C. matruchotii; C .minutissimum; C. mycetoides; C. pilosum; C. pseudodiphtheriticum; "C. pseudogenitalium"; C. pseudotuberculosis; C. renale; CDC groups 1, 2, ANF-1, D-2, E, F-1, F-2, G-1, G-2, and I-2; C. striatum; "C. tuberculostearicum"; C. ulcerans; C. vitarumen; C. xerosis; and Erysipelothrix rhusiopathiae, was typified by significant quantities of hexadecanoic (C16:0) and oleic acids (C18:cis9), with differences in the amounts of linoleic acid (C18:2), stearic acid (C18:0), an unnamed peak (equivalent chain length, 14.966), and small quantities of other known saturated and unsaturated fatty acids. CFA composition of these organisms was sufficiently discriminatory to assist in classification but could not be used as the sole means of identification.

Actinomycetales↗

Description and evaluation of the semiautomated 4-hour rapid ID 32 Strep method for identification of streptococci and members of related genera.

The rapid ID 32 Strep system (bioMérieux, La Balme les Grottes, France) is a new system which allows the identification in 4 h of most streptococci and members of related genera encountered in medical and veterinary bacteriology. Four hundred thirty-three isolates first identified by conventional methods and belonging to the genera Streptococcus, Lactococcus, Enterococcus, Aerococcus, Gemella, Leuconostoc, Erysipelothrix, Gardnerella, and Listeria were tested. Overall, rapid ID 32 Strep correctly identified 413 (95.3%) of the strains, with 109 (25.1%) requiring extra tests for complete identification. Sixteen strains (3.7%) were not identified, and 4 (1.0%) were misidentified. The rapid ID 32 Strep system is a suitable alternative for rapid identification of members of the genus Streptococcus and of related genera.

Bacterial Typing Techniques↗

Alpha-mannosidase: a rapid test for identification of Arcanobacterium haemolyticum.

A 4-h alpha-mannosidase test for identification of Arcanobacterium haemolyticum strains (n = 139) and differentiation of A. haemolyticum from Actinomyces pyogenes strains (n = 30) and other gram-positive rods was evaluated. Practically all A. haemolyticum strains (138 of 139) and the Listeria monocytogenes type strain were alpha-mannosidase positive, while all A. pyogenes strains and Corynebacterium (n = .25) strains as well as the Rhodococcus equi and Erysipelothrix rhusiopathiae type strains were negative. The rapid alpha-mannosidase test, in conjunction with a Gram stain and catalase and reverse CAMP tests, was useful in identification of A. haemolyticum and in differentiation of A. haemolyticum from A. pyogenes and Corynebacterium spp.

Actinomyces↗

Difficulties encountered in identification of a nutritionally deficient streptococcus on the basis of its failure to revert to streptococcal morphology.

Difficulties were encountered in the recognition of a nutritionally deficient streptococcus which continued to display aberrant morphologic forms (especially bulbous swellings and filament formation) despite provision of growth factors. With isolates displaying diverse morphologic entities not characteristic of a given species, e.g., Streptobacillus moniliformis or Erysipelothrix rhusiopathiae, nutritionally deficient streptococci should be considered.

Bacteremia↗

Tiamulin activity against fastidious and nonfastidious veterinary and human bacterial isolates: initial development of in vitro susceptibility test methods.

Tiamulin is a pleuromutilin derivative used in veterinary practice for the control and specific therapy of infections in swine. This report summarizes studies to establish standardized susceptibility testing methods, interpretive criteria, and reagent details for use in veterinary methods recently developed by the National Committee for Clinical Laboratory Standards (NCCLS) (standards M31-A and M37-A, NCCLS, Wayne, Pa., 1999). A total of 636 fastidious and nonfastidious animal and human pathogens were processed by using media and procedures described by the NCCLS. Tiamulin disk diffusion tests used a 30-microg disk concentration, and the proposed MIC breakpoints corresponding to levels achievable in animal target tissues (lung) were < or =4 microg/ml for susceptibility and > or =32 microg/ml for resistance. Correlate zone diameters for specific nonfastidious species were as follows: for Pasteurella multocida and staphylococci tested on Mueller-Hinton agar, susceptibility at > or =19 mm and resistance at < or =11 mm, and for Actinobacillus suis, Erysipelothrix rhusiopathiae, and Streptococcus suis tested on enriched chocolate Mueller-Hinton agar, susceptibility at > or =16 mm and resistance at < or =8 mm. When Actinobacillus pleuropneumoniae was tested, a susceptibility breakpoint of < or =16 microg/ml (> or =9 mm) was suggested for veterinary fastidious medium broth and enriched chocolate Mueller-Hinton agar. Absolute categorical agreement between NCCLS dilution and disk diffusion test results with these criteria ranged from 90.5 to 96.2%. Tiamulin susceptibility testing methods appear to be accurate in their categorical classification for indicated species, and their availability will allow immediate testing of animal isolates to guide therapy via appropriate levels of dosing and to monitor the development of resistance for agents in this unique class.

Humans↗

Enzyme-linked immunosorbent assay employing a recombinant antigen for detection of protective antibody against swine erysipelas.

The specificities and sensitivities of five recombinant proteins of the surface protective antigen (SpaA) of Erysipelothrix rhusiopathiae were examined by indirect enzyme-linked immunosorbent assay (ELISA) with the aim of developing a reliable serological test for the detection of protective antibody against E. rhusiopathiae. Fully mature protein and the N-terminal 416 amino acids (SpaA416) showed sufficient antigenicities, and further examination was done with SpaA416 because of its higher yield. The antibody titers of pigs experimentally immunized with commercial live vaccine and two types of inactivated vaccines clearly increased after immunization, and all pigs were completely protected against challenge with virulent strains. On the other hand, the antibody titers of nonimmunized control pigs remained very low until they were challenged, and all showed severe symptoms or subsequently died. Interference with the production of antibody against live vaccine by maternal antibody or porcine respiratory and reproductive syndrome virus infection 1 week after vaccination was also clearly detected. Because the ELISA titer correlated well with the protection results, the specificity and sensitivity of the ELISA were further evaluated with sera collected from pigs reared on 1 farm on which animals had acute septicemia, 2 farms on which the animals were infected or free from infection, and 10 farms on which the animals were vaccinated with live vaccine, among others. The ELISA titers clearly revealed the conditions of the herds. These results indicate that the SpaA416 ELISA is an effective method not only for evaluating pigs for the presence of protective antibody levels resulting from vaccination or maternal antibody but also for detecting antibody produced by natural infection. This test has important potential for the effective control of swine erysipelas.

Animals↗