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PubMed1982

Is Rhodococcus equi a soil organism?

A total of 189 isolates of Rhodococcus equi and related organisms and 16 marker strains representing the genera Rhodococcus and Corynebacterium were screened for 160 unit characters in a numerical taxonomic study. Analysis of the data indicated that R. equi forms a relatively homogeneous cluster distinctly separated from the recognized species of Rhodococcus and Corynebacterium (sensu stricto). Other members of the genus Rhodococcus are soil organisms and R. equi appears to fit into the genus on ecological as well as taxonomic grounds. It seems unlikely that R. equi could be a gastrointestinal tract commensal because unlike members of the latter group it is an obligate aerobe with an optimum temperature requirement of 28-30 degrees C. It is capable of utilizing simple organic compounds as sources of carbon or carbon and nitrogen and is sensitive to bile salts. Furthermore, it appears that isolation of R. equi from the gut contents of animals is dependent on those animals having access to grazing. Taxonomic studies (backed-up by ecological studies) support the concept that R. equi is a soil organism.

Actinomycetales↗
PubMed2001

Prevalence of virulent Rhodococcus equi in isolates from soil collected from two horse farms in South Africa and restriction fragment length polymorphisms of virulence plasmids in the isolates from infected foals, a dog and a monkey.

The prevalence of virulent Rhodococcus equi in soil isolates from two horse farms in South Africa and nine clinical isolates from six foals, a foal foetus, a dog, and a monkey was investigated. The isolates were tested for the presence of virulence plasmid DNA and 15- to 17-kDa antigens by immunoblotting. Rhodococcus equi was isolated from almost all of the soil samples obtained from the two farms with 5.0 x 10(1) to 3.3 x 10(4) colony forming units per gram of soil. Virulent R. equi was isolated from three soil samples from one of the farms and appeared in 3.8% (three of 80 isolates), but not in any of the 182 isolates from the other farm. Of the three virulent R. equi isolates, one contained an 85-kb type I plasmid and two an 87-kb type I plasmid. Of nine clinical isolates from the foals, foal foetus, dog and monkey, five from the foals were virulent R. equi which expressed the virulence-associated antigens and contained a virulence plasmid 85-kb type I, and were all isolated from cases of pneumonia typical of that induced by R. equi in young foals living in widely separated areas in South Africa. The isolates from the other four foals, the dog and the monkey were avirulent R. equi.

Actinomycetales Infections↗
PubMed1994

Interaction of virulent and non-virulent Rhodococcus equi human isolates with phagocytes, fibroblast- and epithelial-derived cells.

Rhodococcus equi is a facultative, intracellular, Gram-positive coccobacillus, increasingly reported in pneumonia of AIDS-infected patients. We investigated killing resistance properties of human R. equi virulent and avirulent human strains. Avirulent beta-lactam-susceptible strains had lower intracellular colony forming units after 45 min incubation in murine macrophages J774 and human monocyte-macrophage TPH-1 than those of virulent strains. Only virulent beta-lactam-resistant strains persisted within macrophages for at least 18 min only. A beta-lactam-resistant mutant was obtained from a beta-lactam-susceptible strain after selection in a penicillin G-containing culture medium. This mutant strain, like the natural virulent strains, persisted within macrophages, harboured cell-associated appendages, produced phage-like particles and induced, after its intravenous inoculation, a chronic infection in BALB/c nude mice. Supernatant culture of virulent strains transferred partial macrophage-killing resistance properties to avirulent strains. The same supernatant was toxic for L-929, HeLa and Vero cell cultures. These supernatant effects were heat-inactivated, trypsin-inactivated and did not seem to be linked to phage-like particle presence. These data argue that virulence, beta-lactam-resistance, and macrophage-killing resistance are associated in human R. equi isolates. Moreover, only virulent strains produced uncharacterized toxic factors.

Animals↗
PubMed2003

Rapid identification of Rhodococcus equi by a PCR assay targeting the choE gene.

The actinomycete Rhodococcus equi is an important pathogen of horses and an emerging opportunistic pathogen of humans. Identification of R. equi by classical bacteriological techniques is sometimes difficult, and misclassification of an isolate is not uncommon. We report here on a specific PCR assay for the rapid and reliable identification of R. equi. It is based on the amplification of a fragment of the choE gene encoding cholesterol oxidase. The choE-based PCR was assessed by using a panel of strains comprising 132 isolates from different sources and of different geographical origins, all initially identified biochemically as R. equi, and 30 isolates of representative non-R. equi actinomycete species, including cholesterol oxidase producers. The expected 959-bp amplicon was observed only with R. equi isolates, as confirmed by sequencing of a variable region of the 16S RNA gene from a random sample of 20 PCR-positive isolates. All R. equi isolates gave a positive choE-based PCR result, which correlated with a high degree of conservation of the choE gene. Three of the 132 strains originally identified as R. equi were negative for the choE gene, and subsequent analysis of their 16S RNA gene sequences confirmed that they belonged to other bacterial species (Dietzia maris, Mycobacterium peregrinum, and Staphylococcus epidermidis). All non-R. equi isolates were negative by the choE-based PCR. ATCC 21387, the only known isolate of Brevibacterium sterolicum, gave a 959-bp amplicon whose DNA sequence was virtually identical to that of R. equi choE. Comparison of the 16S RNA genes indicated that ATCC 21387 should be considered an R. equi isolate.

Actinomycetales Infections↗
PubMed2004

Hematologic and immunophenotypic factors associated with development of Rhodococcus equi pneumonia of foals at equine breeding farms with endemic infection.

Rhodococcus equi causes severe pyogranulomatous pneumonia in foals and in immunocompromised people. In mice, both CD4+ and CD8+ T lymphocytes contribute to host defense against R. equi, but CD4+ T lymphocytes are required for pulmonary clearance of the bacteria. In this prospective study of 208 foals at two equine breeding farms with endemic R. equi infections, we collected peripheral blood samples at 2 and 4 weeks of age and at the time of diagnosis of R. equi pneumonia. Samples were analyzed for concentrations of total and differential leukocytes, EqCD4+ and EqCD8+ T lymphocytes, and B lymphocytes. Thirty (14.4%) foals developed R. equi pneumonia. At the 2nd week of life, affected foals had significantly lower concentrations of white blood cells (WBC) and segmented neutrophils, significantly lower proportions of EqCD4+ T lymphocytes, and significantly higher proportions of EqCD8+ T lymphocytes. The EqCD4:EqCD8 ratio was significantly lower for affected foals. At the 4th week of life, affected foals had significantly lower concentrations of segmented neutrophils and EqCD4+ T lymphocytes than did unaffected foals. The ratio of EqCD4:EqCD8 was significantly lower for affected foals. Two- and 4-week-old foals with ratios of EqCD4:EqCD8<3 were significantly more likely to develop R. equi pneumonia. There was a significant farm effect which diluted our statistical power to detect differences; however; after adjusting for the farm effect, 2-week-old foals with ratios of EqCD4:EqCD8<3 remained significantly more likely to develop R. equi pneumonia. There were no significant differences in immunophenotypic variables between affected foals (at the time of diagnosis) and age-matched control foals. These data suggest that there are hematologic and immunophenotypic differences between affected and unaffected foals during the first 2-4 weeks of life, prior to onset of clinical signs of R. equi pneumonia. These differences may represent important immunologic mechanisms associated with increased susceptibility of individual foals to infection with R. equi. Because there was considerable overlap between values for affected and unaffected foals, we cannot yet recommend immunophenotyping of foals at endemically-infected farms as a clinically useful screening tool to identify foals at increased risk of developing R. equi pneumonia.

Actinomycetales Infections↗
PubMed2005

Assessment in mice of vapA-DNA vaccination against Rhodococcus equi infection.

There is a need to produce a vaccine against Rhodococcus equi pneumonia in foals in which immunity against infection is largely based on a type 1, cell-mediated, immune response. The VapA protein of the virulence plasmid of R. equi is highly immunogenic. To assess the potential of vapA-DNA to produce immunity, C57BL/6 and BALB/c mice were immunized with a DNA vaccine constructed from vapA incorporated into pcDNA3.1. The plasmid construct expressed VapA in a COS-7 cell line. Intramuscular immunization of mice resulted in enhanced clearance of R. equi from the liver of intravenously challenged mice compared to non-immunized controls. This effect was more marked when pORF-IL-12, a plasmid expressing murine IL12, was included with the vaccine. Antibody developed to VapA, with an IgG2a response being more marked in mice immunized with pcDNA-vapA than in non-immunized or in mice immunized with the mixed vapA and IL-12 plasmid constructs. In conclusion, this study has shown for the first time that DNA immunization with vapA enhances the immune responses of mice against R. equi infection, that the IgG subisotype response is consistent with a type 1-based immune response, and that this can be enhanced by injection of the IL-12 gene.

Actinomycetales Infections↗
PubMed1998

Cytokine induction in murine macrophages infected with virulent and avirulent Rhodococcus equi.

To look for a possible correlation between the virulence of Rhodococcus equi and its cytokine-inducing capacity, we evaluated intracellular survival and measured cytokine induction by mouse macrophages infected with a virulent strain containing an 85-kb plasmid and expressing VapA (103+), its avirulent plasmid-cured derivative (103-), and heat-killed 103+ (HK). After incubation with similar numbers of bacteria, macrophages infected with 103- contained significantly more organisms than those infected with 103+ or HK. The number of bacteria in the macrophages infected with 103- and HK decreased progressively, whereas the 103+ numbers remained constant over 48 h. Interleukin 1beta (IL-1beta), IL-6, IL-10, IL-12 p40, and tumor necrosis factor alpha (TNF-alpha) mRNA induction peaked at 4 h and returned to baseline between 12 and 48 h postinfection. IL-1beta, IL-6, IL-10, and TNF-alpha concentrations assessed by enzyme-linked immunosorbent assay generally agreed well with mRNA expression; IL-12 could, however, not be detected. For all the cytokines detected, mean concentrations in the supernatants were consistently higher in the 103(-)-infected monolayers than in those infected with 103+, although, with the exception of IL-1beta, the differences were not statistically significant. R. equi HK was a poor inducer of cytokine production. In conclusion, virulent and avirulent R. equi strains induced similar levels of cytokine synthesis. The slightly greater induction of most cytokines observed following infection with 103- is likely secondary to greater uptake by macrophages rather than to a direct role of VapA or another plasmid-encoded product in downregulating cytokine induction.

Animals↗
PubMed2002

Cholesterol oxidase and resistance of Rhodococcus equi to peroxidative stress in vitro in the presence of cholesterol.

Rhodococcus equi is a well-characterized bacterial pathogen which lyses cell membranes with the help of cholesterol oxidase (CO). Survival in macrophages is warranted by its ability to resist reactive radicals via catalase and superoxide dismutase (SOD). Therefore, CO production in the absence or presence of 0.1 % cholesterol and sensitivity to exogenous hydrogen peroxide (H2O2) and superoxide anion (SOA) were tested in seven strains of R. equi in vitro. When R. equi strains were grown on agar plates with cholesterol, the bacterial growth [colony-forming units (cfu)/plate] did not increase significantly in comparison with the growth on plates without cholesterol. The activity of CO increased, significantly for extracellular CO. In subsequent experiments, R. equi strains grown on cholesterol were stressed with H2O2 or SOA so that approximately 10 % of cfu/plate survived. During stress induced by SOA, membrane CO and SOD activity increased significantly. Catalase activity increased 2-fold with H2O2 and 3-fold with SOA exposure. These data suggest that the presence of cholesterol induces CO in bacteria grown on agar plates. Catalase, SOD and even membrane-bound CO respond to reactive oxygen species.

Actinomycetales Infections↗
PubMed2000

Evaluation of nasotracheal aspiration as a diagnostic tool for Rhodococcus equi pneumonia in foals.

The reliability of preparing bacteriological cultures from nasotracheal aspirates of foals routinely in order to diagnose R. equi pneumonia in foals was studied by isolating Rhodococcus equi from specimens obtained from 96 foals by nasotracheal aspiration with a silicon catheter. Results were compared with specimens obtained from 21 foals by transtracheal aspiration (percutaneous tracheal puncture). These 117 foals showed clinical signs of respiratory tract infection at sampling. R. equi was isolated from 14 of 21 (66.7%) specimens by transtracheal aspiration and from 59 of 96 (61.4%) specimens by nasotracheal aspiration, 649 of 655 isolates (99.1%) from the 73 positive specimens were virulent R. equi, and the culture-positive foals were diagnosed as having R. equi pneumonia. To assess the contamination of aspirates by organisms from the nasopharynx, the results of R. equi isolation from nasal swabs obtained from 56 of the 96 foals were compared to those obtained by nasotracheal aspiration from the same foals. R. equi was isolated from 2 of the 56 nasal swabs: one from a tracheal aspirate was positive, and the other was not. These results suggest that the nasotracheal aspiration technique, which is noninvasive and not associated with complications, could be used as an alternative to the transtracheal aspiration method, especially for the diagnosis of R. equi pneumonia in foals.

Actinomycetales Infections↗
PubMed1992

Virulence-associated 15- to 17-kilodalton antigens in Rhodococcus equi: temperature-dependent expression and location of the antigens.

Virulent Rhodococcus equi showing 15- to 17-kDa antigens, which is virulent in mice, was found to harbor an 85-kbp plasmid, and the 15- to 17-kDa antigens were found to be associated with possession of the 85-kbp plasmid of R. equi (S. Takai, T. Sekizaki, T. Ozawa, T. Sugawara, Y. Watanabe, and S. Tsubaki, Infect. Immun. 59:4056-4060, 1991). The expression of these antigens was temperature regulated: when cells were grown at a low temperature (25 to 32 degrees C), they did not express them, whereas they expressed them in large amounts when the cells were grown at a higher temperature (34 to 41 degrees C). The antigens were expressed on the cell surface, as evidenced by their susceptibility to proteolysis by a trypsin and by the biotin-avidin protein-blotting technique.

Antigens, Bacterial↗
PubMed1986

Quantitative fecal culture for early diagnosis of Corynebacterium (Rhodococcus) equi enteritis in foals.

Quantitative culture of Corynebacterium (Rhodococcus) equi from feces of 17 foals on a farm (A) with an endemic C. equi infection problem and 26 foals on a farm (B) without the disease in the past decade was done with a selective medium at weekly or monthly intervals from April to August of 1984. Corynebacterium equi was observed in the feces of 16 of 17 foals on farm A, and 19 of 26 foals on farm B. The mean viable count of C. equi in one gram of feces was 4.1 +/- 3.7 (log10) on farm A, and 3.9 +/- 3.4 (log10) on farm B. Corynebacterium equi was recovered from feces of foals as young as two weeks old. Almost all foals at an age between two to four weeks shed the bacteria in the feces. During the observation period two foals showed clinical signs: fever, diarrhea, and cough, at four or five weeks old. At the same time the bacterial count per gram of feces increased from 4 to 7 or 8 (log10). They shed large number of bacteria in the feces and continued to show the clinical signs until death at 10 or 11 weeks old. One of the foals was diagnosed as having had C. equi enteritis and pneumonia by the postmortem recognition of lesions with bacteriological confirmation. The quantitative culture of the feces of foals at weekly intervals after birth on farm A was found to be very useful as an aid in early diagnosis of C. equi enteritis in foals.

Actinomycetales Infections↗
PubMed1996

Rhodococcus equi infection in 3 AIDS patients.

Three cases of AIDS complicated by Rhodococcus equi infection are reported. At least one of the patients acquired his Rhodococcus infection in Africa. Despite the fact that the R. equi strains were susceptible to tetracycline, erythromycin, amikacin, co-trimoxazole, rifampicin and vancomycin, these antibiotics were clinically not successful. A clinical improvement was observed in only one patient during teicoplanin and imipenem-cilastatin treatment. Multicentre clinical trials are needed to determine the optimal treatment of R. equi infections in AIDS patients.

AIDS-Related Opportunistic Infections↗
PubMed1998

Physical and serologic examinations of foals at 30 and 45 days of age for early diagnosis of Rhodococcus equi infection on endemically infected farms.

OBJECTIVE: To evaluate results of physical and serologic examinations of foals at 30 and 45 days of age on 3 types of farms with various prevalences of clinical disease (endemic, sporadic, none) caused by Rhodococcus equi and to determine whether evaluations were helpful in early diagnosis and control of the disease. DESIGN: Prospective cohort study. ANIMALS: 144 foals at 30 and 45 days of age. PROCEDURE: During a 2-year period, 36 foals on farms at which R equi infection was endemic, 71 foals on farms at which the disease was sporadically detected, and 37 foals on farms without the disease were examined by means of auscultation of lungs, serum biochemical and hematologic analyses, and determination of antibody titers against R equi, using ELISA. Transtracheal aspirates were obtained from 14 of 32 foals that had clinical signs of disease and 7 of 41 seropositive foals that did not have clinical signs of disease. RESULTS: Prevalences of respiratory tract disease and seropositive conversion rates for 45-day-old foals on endemically and sporadically infected farms were significantly higher than on farms without the disease. Rhodococcus equi was isolated from tracheal aspirates of seropositive foals, even when clinical signs were not evident. CLINICAL IMPLICATIONS: Physical and serologic examinations of foals at 30 and 45 days of age were useful for early diagnosis of R equi infection, especially for foals on farms at which the disease was endemic.

Actinomycetales Infections↗
PubMed1989

Rhodococcus equi: an emerging opportunistic pathogen?

Human infection with Rhodococcus equi is apparently rare with most published reports describing the development of lung abscesses in immunocompromised hosts. Of only 18 cases of infection previously recorded, four have recently occurred in patients with the acquired immune deficiency syndrome (AIDS). In Australasia, R. equi has frequently been isolated from soil and infected farm animals yet no human infections have been reported thus far. Three cases of R. equi infection have occurred in New Zealand and, collectively, they cover a wider spectrum of disease than that previously recognised. The natural history of R. equi infections, their clinical features and treatment are described in the light of our recent experience.

Acquired Immunodeficiency Syndrome↗
PubMed1991

Mycolic acid-containing glycolipid as a possible virulence factor of Rhodococcus equi for mice.

By the use of various Rhodococcus equi strains differing in the length of carbon chains of glycolipid, we examined whether the glycolipid, glucose monomycolate, was contributing to the virulence of R. equi for mice. R. equi strains with longer carbon chain mycolic acid showed a higher virulence as determined by lethality and granuloma formation in mice than those with shorter ones. When purified glycolipid was injected into mice, granuloma formation and liver damage were most prominent with the glycolipid having longer carbon chain mycolic acid. Only a representative strain with longer carbon chain mycolic acid persisted in the spleen of mice after intravenous injection, while a strain with shorter carbon chain mycolic acid was readily eliminated. These results suggested that glycolipid was at least one of the virulence factors of R. equi and that the carbon chain length of mycolic acid might be critical in the expression of virulence.

Actinomycetales Infections↗
PubMed1987

The ecology of Rhodococcus equi and physicochemical influences on growth.

Growth of Rhodococcus equi was studied in vitro. Optimal growth occurred under aerobic conditions between pH 7.0 and 8.5, at 30 degrees C. R. equi survived better in a neutral soil (pH 7.3) than it did in two acid soils (pH less than 5.5). It grew substantially better in soils enriched with faeces than in soils alone. Simple organic acids in horse dung, especially acetate and propionate, appear to be important in supporting growth of R. equi in the environment. The ecology of R. equi can be best explained by an environmental cycle allowing its proliferation in dung, influenced by management, grazing behaviour and prevailing climatic conditions. Preventive measures should be aimed at reducing or avoiding focal areas of faecal contamination in the environment.

Animals↗
PubMed2003

Rhodococcus equi and genetic susceptibility: assessing transferrin genotypes from paraffin-embedded tissues.

Rhodococcus equi is a bacterial pathogen, ubiquitous in the soil, that infects many foals and is lethal to some. Transferrin is an iron-binding protein that has bacteriostatic properties in the blood. Transferrin is also highly polymorphic in most species, with 15 variants identified for horses using biochemical methods, and may be responsible for variation in susceptibility to bacterial pathogens. The objectives of this study were 1) to compare biochemical typing with DNA typing of transferrin, 2) to determine if transferrin DNA from archival paraffin-embedded tissue samples can be recovered and typed with molecular methods, and 3) to determine if there was an association between foal death caused by R. equi and transferrin type. Comparing biochemical methods and DNA sequencing for 41 horses demonstrated correspondence between the typing methods. The allele frequency of archival paraffin-embedded tissue samples from 34 Thoroughbred foals that succumbed to R. equi showed an excess of the F allele and a deficiency of the D allele (P < 0.05). Year of collection (P > 0.8), age of foal (P > 0.3), and sex of foal (P > 0.6) were not statistically associated with transferrin type. The archival material was successfully transferrin typed using DNA sequencing, and there may be an association between foal death caused by R. equi and transferrin type.

Actinomycetales Infections↗
PubMed2005

Protective role of neutrophils in mice experimentally infected with Rhodococcus equi.

Neutrophils are important in controlling early infections with the intracellular bacterium Rhodococcus equi. Antineutrophil monoclonal antibody (RB6-8C5)-induced neutrophil deficiency during the first week after experimental infection of mice with R. equi resulted in more severe disease and significantly increased tissue concentrations of R. equi.

Actinomycetales Infections↗