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Rhodococcus equi emerged as a zoonotic pathogen of human immunodeficiency virus-infected patients over the last three decades. Two virulence plasmid types of R. equi, pVAPA and pVAPB associated with equine and porcine isolates, have been recognized, and more recently, pVAPN, a novel host-associated virulence plasmid in R. equi, was found in bovine and caprine isolates. We reinvestigated 39 previously reported isolates of R. equi from patients with and without acquired immunodeficiency syndrome (AIDS) by detecting vapA, vapB and vapN using PCR and plasmid profiling. After excluding one isolate that could not be cultured from frozen storage, eight isolates carried a virulence plasmid encoding vapA (pVAPA), 10 carried a virulence plasmid encoding vapB (pVAPB), seven carried a virulence plasmid encoding vapN (pVAPN) and 13 were negative for those genes. Of the 29 isolates from patients with AIDS, 7, 10 and 5 harboured pVAPA, pVAPB and pVAPN respectively. Among nine isolates from patients without AIDS, one and two harboured pVAPA and pVAPN respectively. This study demonstrated that pVAPN-positive R. equi existed in human isolates before 1994 and reaffirmed that equine-associated pVAPA-positive, porcine-associated pVAPB-positive and bovine- or caprine-associated pVAPN-positive R. equi are widely spread globally. Because domestic animals might be major sources of human infection, further research is needed to reveal the prevalence of pVAPN-positive R. equi infection in cattle and goats., (© 2020 The Society for Applied Microbiology.)

A recent taxonomic study confirmed the synonymy of Rhodococcus equi (Magnusson 1923) Goodfellow and Alderson 1977 and Corynebacterium hoagii (Morse 1912) Eberson 1918. As a result, both R. equi and C. hoagii were reclassified as Rhodococcus hoagii comb. nov. in application of the principle of priority of the Prokaryotic Code. Because R. equi is a well-known animal and zoonotic human pathogen, and a bacterial name solidly established in the veterinary and medical literature, we and others argued that the nomenclatural change may cause error and confusion and be potentially perilous. We have now additionally found that the nomenclatural type of the basonym C. hoagii , ATCC 7005 T , does not correspond with the original description of the species C. hoagii in the early literature. Its inclusion as the C. hoagii type on the Approved Lists 1980 results in a change in the characters of the taxon and in C. hoagii designating two different bacteria. Moreover, ATCC 7005, the only strain in circulation under the name C. hoagii , does not have a well documented history; it is unclear why it was deposited as C. hoagii and a possible mix-up with a Corynebacterium ( Rhodococcus ) equi isolate is a reasonable assumption. We therefore request the rejection of Rhodococcus hoagii as a nomen ambiguum , nomen dubium and nomen perplexum in addition to nomen periculosum , and conservation of the name Rhodococcus equi , according to Rules 56ab of the Code.

Chitin deacetylase (CDA) can hydrolyse the acetamido group of chitin polymers to produce chitosans, which are used in various fields including the biomedical and pharmaceutical industries, food production, agriculture, and water treatment. CDA represents a more environmentally-friendly and easier to control alternative to the chemical methods currently utilised to produce chitosans from chitin; however, the majority of identified CDAs display activity toward low-molecular-weight oligomers and are essentially inactive toward polymeric chitin or chitosans. Therefore, it is important to identify novel CDAs with activity toward polymeric chitin and chitosans. In this study, we isolated the bacterium Rhodococcus equi F6 from a soil sample and showed that it expresses a novel CDA (ReCDA), whose activity toward 4-nitroacetanilide reached 19.20 U/mL/h during fermentation and was able to deacetylate polymeric chitin, colloidal chitin, glycol-chitin, and chitosan. Whole genome sequencing revealed that ReCDA is unique to the R. equi F6 genome, while phylogenetic analysis indicated that ReCDA is evolutionarily distant from other CDAs. In conclusion, ReCDA isolated from the R. equi F6 strain expands the known repertoire of CDAs and could be used to deacetylate polymeric chitosans and chitin in industrial applications.

Background: Rhodococcus equi (R. equi) infections are endemic in many horse facilities in the United States resulting significant economic loses annually. Currently, there is no commercial vaccine available and the emergence of isolates that are resistant to the current treatment and prophylaxis using antibiotics prompts closer surveillance of this pathogen., Objective: This study compares three different genotyping techniques, Pulsed Field Gel Electrophoresis (PFGE), Multilocus Sequence Typing (MLST) and whole genome SNP-based phylogeny to determine the most accurate method to monitor the spread of macrolide-and-rifampin-resistant R. equi., Methods: 16 macrolide and rifampin-resistant and 6 susceptible R. equi and their Illumina Miseq whole genome sequences were used in this study. The isolates were sub-typed by PFGE with VspI and a dendrogram based on their similarities generated. Additionally, three phylogenetic trees were constructed using CSI phylogeny on (i) whole genome sequences (WGS), (ii) in silico MLST sequences and (iii) MLST sequences obtained after PCR-amplification and Sanger sequencing., Results: PFGE identified 18 different genetic profiles and grouped the 22 isolates into 3 clusters independently of their susceptibilities. The phylogenetic trees built from WGS and MLST data showed similar topology, separating the isolates into 2 major clades in accordance with their susceptibility profiles (susceptible and resistant). However, only the trees generated with next generation sequencing data could detect the clonality of the resistant isolates., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Objective: Rhodococcus equi is an opportunistic pathogen that causes disease worldwide in young foals and immunocompromised humans. The interactions of R. equi with the host immune system have been described; however, most studies have been conducted using a few well-characterized strains. Because biological differences between R. equi strains are not well characterized, it is unknown if experimental results will replicate when different strains are used. Therefore, our objective was to compare the growth and biofilm formation of low-passage-rate clinical isolates of R. equi to higher-passage-rate, commonly studied isolates to determine whether strain-to-strain variation exists., Results: Twelve strains were used: 103+, ATCC 33701, UKVDL206 103S harboring a GFP-expressing plasmid, a plasmid-cured 33701 (higher-passage-rate) and seven low-passage clinical isolates. Generation time in liquid revealed fast, moderate-fast, moderate-slow, and slow-growing isolates. The higher-passage-rate isolates were among the moderate-slow growing strains. A strain's rate of growth did not correspond to its ability to form biofilm nor to its colony size on solid media. Based on our results, care should be taken not to extrapolate in vitro work that may be conducted using different R. equi strains. Further work is needed to evaluate the effect that the observed differences may have on experimental results.

Rhodococcus equi causes pulmonary and extrapulmonary infections in animals and humans, with endemic situations and significant young foal mortality in stud farms worldwide. Despite its economic impact in the horse-breeding industry, the broad geographic and host distribution, global diversity and population structure of R. equi remain poorly characterised. In this context, we developed a multilocus sequence typing (MLST) scheme using 89 clinical and environmental R. equi of various origins and eight Rhodococcus sp. Data can be accessed at http://pubmlst.org/rhodococcus/. A clonal R. equi population was observed with 16 out of 37 sequence types (STs) grouped into six clonal complexes (CC) based on single-locus variants. One of the six CCs (CC3) is not host-specific, suggesting potential exchanges between different R. equi reservoirs. Most of the virulent equine R. equi CCs/unlinked STs were plasmid-type-specific. Despite this, marked genetic variability with the circulation of multiple R. equi genotypes was generally observed even within the same animal. Focusing on outbreaks, data indicated (i) the potential contagious transmission of R. equi during the 2012-Mayotte equine outbreak because of the poor genotype diversity of clinical strains; (ii) a potential porcine outbreak among the 30 Belgian farms investigated in 2013. This first Rhodococcus equi MLST is a powerful tool for further epidemiological investigations and population biology studies of R. equi isolates., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Background: Rhodococcus equi is an emerging zoonotic presumably foodborne pathogen. Since the data on the worldwide prevalence of R. equi in meat animals are scarce, the present study aimed to investigate the molecular epidemiology of R. equi in swine, cattle and horse carcasses intended for human consumption in Poland., Results: Totally 1028 lymph node samples were examined. R. equi was isolated from 26.6 % (105/395) swine and 1.3 % (3/234) bovine healthy submaxillary lymph nodes. In horses, R. equi was isolated only from 0.5 % (1/198) samples of middle tracheo-branchiales lymph node while no lymphocentrum retropharyngeum sample was positive (0/198). The purulent lesions were observed only in 0.8 % swine submaxillary lymph nodes samples (3/398) and in two of them R. equi was detected. All bovine and most of swine isolates (98.1 %) were vapB-positive. 87.9 % of swine isolates carried 95-kb type 5 plasmid, 3.7 % type 1 and plasmid types: 4, 7, 10, 11, 21, 31 were carried by a single isolate (0.9 %). All bovine isolates carried VAPB type 26. Single horse isolate was vapA-positive and carried plasmid VAPA 85-kb type I., Conclusions: The prevalence of vapB-positive R. equi in investigated healthy swine intended for human consumption was very high. Not only swine, but also even apparently healthy cattle or horse carcasses should be considered as a potential source of R. equi for humans, especially in countries where undercooked or raw beef or horsemeat is traditionally consumed.

Rhodococcus defluvii strain Ca11(T) was isolated from a bioreactor involved in extensive phosphorus removal. We have sequenced the whole genome of this strain, and our comparative genomic and phylogenetic analyses confirm its close relatedness with Rhodococcus equi (Rhodococcus hoagii) strains, which share >80 % of the gene content. The R. equi virulence plasmid is absent though most of the chromosomal R. equi virulence-associated genes are present in R. defluvii Ca11(T). These data suggest that although R. defluvii is an environmental organism, it has the potential to colonize animal hosts.

Rhodococcus equi is a soil saprophyte and an opportunistic pathogen causing infections in animals, and rarely in humans. The presence of R. equi in tissues and faeces of some wild animal species was demonstrated previously. In this study we characterized R. equi isolates from submaxillary lymph nodes of free-living wild boars (n=23), red deer (n=2) and roe deer (n=2). This is the first description of R. equi strains isolated from tissues of the Cervidae. All isolates were initially recognized as R. equi based on the phenotypic properties. Their identification was confirmed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, detection of the choE gene and by sequence analysis of the 16S rRNA and rpoB genes. The presence of three plasmidic genes (traA, vapA and vapB) associated with R. equi virulence was investigated by PCR. In 16 wild boar isolates the traA and vapB genes were detected and they were located on virulence plasmids type 5, 7 or 11. The isolates from cervids and the remaining wild boar isolates were classified as avirulent based on a genotype traA(-)/vapA(-)B(-). In summary, these results confirm that wild boars can be a source of intermediately virulent R. equi strains, and indicate that red deer and roe deer can be a reservoir of avirulent R. equi strains., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Equine infection with Corynebacterium pseudotuberculosis can manifest in several forms, including external or internal abscesses. The objective of this study was to phenotype clinical isolates of C. pseudotuberculosis and to investigate the relationship between lesion location and extent of lesions in the animals from which they were collected. One hundred and seventy-one C. pseudotuberculosis biovar equi isolates were collected from horses presenting to the University of California Veterinary Medical Teaching Hospital and two other sources in the period between September 1996 and December 2011. Bacterial isolates were grouped on the bases of biochemical characteristics and growth on brain heart infusion agar. Six phenotypes were identified: (1) large colonies that metabolized sucrose (n = 81); (2) large sucrose-negative colonies (n = 47); (3) medium sucrose-positive (n = 20); (4) medium sucrose-negative (n = 11); (5) small sucrose-positive (n = 7), and (6) small sucrose-negative (n = 5). Medical records corresponding to each isolate were accessed from the University's administrative computer system or from the submitting source in order to determine the anatomical site from which the isolate was collected (n = 171), as well as the extent of lesions (n = 164) in the patient. The relationship between phenotype, lesion location and extent of lesions was then investigated statistically. No significant relationship between strain and lesion location or extent of lesions was found. This suggests that phenotypic differences during in vitro culture does not account for external versus internal disease in horses. Further work to characterize strains genotypically and to identify determinants for bacterial virulence should be performed. Importantly, host and environmental factors should also be further investigated., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

A recent review of the nomenclatural history of Rhodococcus equi and its heterotypic synonyms reveals a situation in which the strict application of the Rules of the International Code of Nomenclature of Prokaryotes have resulted in the renaming of this known zoonotic pathogen, which may be reasonably viewed as a perilous name. This situation can be remedied only by the Judicial Commission rendering an opinion to conserve the name Rhodococcus equi and to reject its earlier heterotypic synonym, Corynebacterium hoagii.

Based on a nomenclatural point of view, the name Rhodococcus equi is associated, as required by the Bacteriological Code, with a defined position, rank and circumscription. A search of the literature indicates that the name Rhodococcus equi (Magnusson 1923) Goodfellow and Alderson 1977 has also been shown to be a synonym of Corynebacterium equi Magnusson 1923, Corynebacterium hoagii (Morse 1912) Eberson 1918 and Nocardia restricta (Turfitt 1944) McClung 1974. Application of the rules of the Bacteriological Code together with the currently inferred taxonomic concept associated with the species bearing the name Rhodococcus equi indicates that this is not the correct name of this taxon and the use of that name in the context of a circumscription that includes the type strain of the species Corynebacterium hoagii is contrary to the Rules of the Code.

The taxonomic status of Rhodococcus equi, originally isolated from foal specimens, has been the subject of discussion for a number of years. The chequered history of the taxon has prompted this polyphasic analysis of R. equi strains, close members of the genus Rhodococcus and representatives of other genera classified in the order Corynebacteriales, to establish the taxonomic position of this taxon. Thirty one R. equi strains, including the type strain, were examined for genotypic and numerical taxonomic properties. The resultant data are consistent with their classification in the order Corynebacteriales but the R. equi strains formed a distinct phyletic clade away from representatives of other members of the genus Rhodococcus in the 16S rRNA gene tree. Representatives of this clade shared their highest pairwise 16S rRNA gene sequence similarities with the type strain of Rhodococcus kunmingensis (95.2-98.1 %). However, the R. equi taxon was readily distinguished from R. kunmingensis and from the other members of the order Corynebacteriales using a combination of genotypic, chemotypic and phenotypic properties. On the basis of these data the R. equi strains are considered to represent a new genus. The name proposed for this taxon is Prescottia gen. nov., with Prescottia equi comb. nov. as the type species containing the type strain, C 7(T) (= ATCC 25729(T) = ATCC 6939(T) = CCUG 892(T) = CIP 54.72(T) = DSM 20307(T) = HAMBI 2061(T) = NBRC 14956(T) = JCM 1311(T) = JCM 3209(T) = LMG 18452(T) = NBRC 101255(T) = NCTC 1621(T) = NRRL B-16538(T) = VKM Ac-953(T)).

Malacoplakia is a rare inflammatory condition characterized by the accumulation of benign macrophages associated with pathognomonic Michaelis-Gutmann bodies (MGBs). It is usually found in the genito-urinary tract, and has been associated with immunocompromised states. In this short report, we present 5 patients with pulmonary nodules clinically suspicious for primary or metastatic lung cancer. The histologic examination of the surgical specimens revealed a nonspecific granulomatous chronic disease, and despite the paucity of classical MGBs, a pulmonary malacoplakia was suspected. In all cases the opportunistic pathogen Rhodococcus equi (R. equi) was identified by 16S rRNA gene sequence analysis, leading to the final pathological diagnosis of malacoplakia. We conclude that pulmonary malacoplakia associated with R. equi is a rare disease affecting also immunocompetent patients. The pathogenesis and the diagnostic problems are discussed. Since infection by R. equi is treatable, the importance of its early recognition should be emphasized.

The identification of 83 Corynebacterium, 13 Arcanobacterium haemolyticum, and 10 Rhodococcus equi strains by conventional methods (API Coryne complemented with 16S rRNA gene sequence analysis) was compared with matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry identification. The correlation between API and MALDI-TOF results was 89%. MALDI-TOF is a rapid and accurate system for identification of the above-mentioned microorganisms.

To characterize the potential epidemiological relationship between the origin of Rhodococcus equi strains and the type of their virulence plasmids, we performed a comparative analysis of virulence plasmid types encountered in 96 R. equi strains isolated from (1) autopsied horses, (2) organic samples (horse faeces, manure and straw) and (3) environmental samples. Our results revealed no clear epidemiological link between virulence plasmid type and the origin of R. equi strains isolated from horse-related environments. To understand this result, we determined the nucleotide sequence of the second most frequently isolated virulence plasmid type: a 87-kb type I (pVAPA116) plasmid and compared it with the previously sequenced (and most commonly encountered) 85-kb type I (pVAPA1037) plasmid. Our results show that the divergence between these two plasmids is mainly due to the presence of three allelic exchange loci, resulting in the deletion of two genes and the insertion of three genes in pVAPA116 compared with pVAPA1037. In conclusion, it appears that the divergence between the two sequenced rhodococcal virulence plasmids is not associated with the vap pathogenicity island and may result from an evolutionary process driven by a mobility-related invertase/resolvase invA-like gene., (© 2010 ANSES. Journal compilation © 2010 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd.)

The mutant prevention concentration (MPC) for ciprofloxacin was determined for two Rhodococcus equi strains. The MPC for both strains was 32 mug/ml, which is above the peak serum concentration of ciprofloxacin obtainable by oral administration in humans. Nine single nucleotide changes corresponding to eight amino acid substitutions in the quinolone resistance-determining regions of DNA gyrase subunits A and B were characterized. Only mutants with amino acid changes in Ser-83 of GyrA were highly resistant (>or=64 microg/ml). Our results suggest that ciprofloxacin monotherapy against R. equi infection may result in the emergence of ciprofloxacin-resistant mutants.

During January to April 2007, hospital staff reported 3 patients with Rhodococcus equi bloodstream infections. Isolates were analyzed at the Centers for Disease Control and Prevention, Atlanta, GA, to confirm identification and to assess strain relatedness; 2 were R. equi but genetically distinct, and 1 was identified as Gordonia polyisoprenivorans. Rapid reference laboratory support prevented an unnecessary outbreak investigation., ((c) 2010 Elsevier Inc. All rights reserved.)

Two hundred and twelve Rhodococcus equi strains were isolated from soil, nasal and rectal swabs of horses and immunocompromised human patients in Hungary and serotyped using Prescott's serotyping system. One hundred and forty-seven strains (69.3%) belonged to serotype 1, 22 strains (10.4%) to serotype 2, 6 strains (2.8%) to serotype 3 and 1 strain (0.5%) to serotype 4. Serotypes 5, 6 and 7 were not found and 36 strains (17%) could not be typed. Serotype 1 (72%) was the type most commonly isolated from clinical samples of foals or from the soil of horse facilities. Six out of 8 R. equi strains from humans belonged to serotype 2, and two human strains were untypable. The data show that the prevalence of R. equi serotypes varies in different geographic areas of the country.

The plasmid profiles of virulent Rhodococcus equi strains isolated on three horse-breeding farms located in different parts of Hungary were investigated. From 49 soil samples collected on the three farms, 490 R. equi isolates (10 from each sample) were obtained and tested for the presence of 15- to 17-kDa antigens (VapA) by immunoblotting and PCR. Ninety-eight VapA-positive isolates were detected from 30 of the 49 culture-positive samples with a prevalence ranging from 13.1% to 23.2%. Of the 98 virulent isolates, 70 contained an 85-kb type I plasmid, 13 contained an 87-kb type I plasmid, and 15 contained an 85-kb type III plasmid which had been uniquely isolated from soil isolates in the United States. This study demonstrates that the virulent form of R. equi is very widespread in the soil environment of these stud farms in Hungary and the plasmid pattern is different from farm to farm.

The plasmid types and serotypes of 164 Rhodococcus equi strains obtained from submaxillary lymph nodes of swine from different piggeries in 28 villages and towns located throughout the country were examined. The strains were tested by PCR for the presence of 15- to 17-kDa virulence-associated protein antigen (VapA) and 20-kDa virulence-associated protein antigen (VapB) genes. Plasmid DNAs were isolated and analyzed by digestion with restriction endonucleases to estimate size and compare their polymorphism characteristics. None of the 164 isolates contained the vapA gene, and 44 (26.8%) isolates were positive for the vapB gene, showing a product of the expected 827-bp size in the PCR amplification. The 44 isolates of intermediate virulence contained virulence plasmids that were identified as types 1 (3 isolates), 4 (1 isolate), 5 (36 isolates), 6 (1 isolate), and 7 (2 isolates) and as a new variant (1 isolate). On the basis of restriction digestion patterns of plasmid DNAs, we tentatively designated the variant as type 17. Use of the serotyping method of Prescott showed that 110 (67.1%) out of the 164 isolates were typeable and that serotype 2 predominated (83 isolates [50.6%]), followed by serotype 1 (26 strains [15.9%]). Only one isolate belonged to serotype 3. A total of 54 (32.9%) isolates were untypeable in Prescott's system. The prevalence of R. equi strains of intermediate virulence among the isolates that came from the submaxillary lymph nodes of swine in Hungary was lower than that seen with isolates obtained elsewhere.

A case of pyogenic liver abscess (PLA) due to Rhodococcus equi in an immunocompetent individual was successfully treated by combining surgery and antibiotics. The R. equi-targeted antimicrobial agents erythromycin and rifampin were used only after surgical resection of the lesion and identification of the infective organism.

We have isolated four strains of Rhodococcus which specifically degrade estrogens by using enrichment culture of activated sludge from wastewater treatment plants. Strain Y 50158, identified as Rhodococcus zopfii, completely and rapidly degraded 100 mg of 17beta-estradiol, estrone, estriol, and ethinyl estradiol/liter, as demonstrated by thin-layer chromatography and gas chromatography-mass spectrometry analyses. Strains Y 50155, Y 50156, and Y 50157, identified as Rhodococcus equi, showed degradation activities comparable with that of Y 50158. Using the random amplified polymorphism DNA fingerprinting test, these three strains were confirmed to have been derived from different sources. R. zopfii Y 50158, which showed the highest activity among these four strains, revealed that the strain selectively degraded 17beta-estradiol during jar fermentation, even when glucose was used as a readily utilizable carbon source in the culture medium. Measurement of estrogenic activities with human breast cancer-derived MVLN cells showed that these four strains each degraded 100 mg of 17beta-estradiol/liter to 1/100 of the specific activity level after 24 h. It is thus suggested that these strains degrade 17beta-estradiol into substances without estrogenic activity.

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.

Rhodococcus equi is an opportunistic pathogen that usually causes infection in immunocompromised hosts. A heart transplant recipient who had been treated with amphotericin B for pulmonary aspergillosis showed newly developed multiple nodules with a central necrotic area in the right lower lobes. Cultures of several blood samples and an aspirate of the lung nodule yielded a Gram-positive coccobacillary bacterium, which was initially reported as a Corynebacterium species, but was later identified as R. equi by API CORYNE (bioMerieux SA, Marcy l'Etoile, France) and by demonstrating the production of 'equi factor'. The identification was subsequently confirmed by an R. equi-specific polymerase chain reaction (PCR). The patient was successfully treated with ciprofloxacin and azithromycin for 14 weeks. This is the first documented case of R. equi infection in Korea. There is a possibility of underestimation of R. equi infections due to the misidentification of the organism as a contaminating diphtheroid. Because R. equi will not respond to the conventional empirical therapy, the microbiology laboratory should identify R. equi in a timely manner. R. equi-specific PCR will be a useful confirmatory test in human infection.

Rhodococcus equi isolates (204) obtained from foals (lung abscesses, lymph nodes, nasal discharge, rectal swabs) bred in 15 studs located throughout Hungary, isolates from soil samples, lymph nodes of pigs and from lesions of human patients were examined to determine genotypic diversity of virulence-associated plasmids. Isolates were tested for the presence of 15-17 kDa virulence-associated protein antigen (VapA) and 20k Da (VapB) genes by polymerase chain reaction (PCR). Plasmid DNAs were isolated and analysed by digestion with restriction endonucleases for estimation of size and comparison of polymorphisms. Of 146 clinical isolates from foals in 15 studs, 129 (88.3%) gave positive results for the VapA gene, showing a 564 bp product of the expected size in the PCR amplification. Of the 129 clinical isolates from foals, 123 contained an 85 kb type I plasmid and the remaining six contained an 87 kb type I plasmid. Of 48 soil isolates from two horse studs, 26 (54.2%) were positive for VapA gene and contained an 85 kb type I plasmid. Of three pig isolates, one was positive for VapA gene and contained an 85 kb type I plasmid, and the remaining two were positive for the VapB gene, showing a 827 bp product of the expected size in the PCR amplification and were R. equi of intermediate virulence which contained a 95 kb type S5 plasmid. Of the seven human isolates, five were positive for VapB gene by PCR, these were R. equi of intermediate virulence, which contained a 95 kb type S5 plasmid. These results revealed that virulent R. equi strains harbouring a virulence plasmid of 85 kb type I or 87 kb type I, which have been found in clinical isolates from Europe and North and South America, are widespread in Hungary. Furthermore, same intermediately virulence plasmid type was found in both human and pig isolates.

Pulsed-field gel electrophoresis of restriction endonuclease-digested genomic DNA from a large collection of clinical isolates of Rhodococcus equi, an important pathogen of foals, was used to compare strain distribution between farms and over time. Forty-four strains were found among 209 isolates, with 5 of these accounting for over half the isolates and the 22 strains isolated more than once accounting for 90% of the isolates. The average genotypic diversity on each farm and in each year was found to be less than the genotypic diversity of the isolates taken as a whole, with 5.2% of the total diversity being due to differences between farms and 5.5% to differences between years. A small number of strains on each farm were found to have caused at least half the clinical cases of disease, and these varied between farms and, to a lesser extent, years. Most strains were found on more than one farm, and some very similar restriction patterns were found among isolates from different continents, indicating that strains can be very widespread. Multiple strains were isolated in five of the six cases in which more than one isolate from a single foal was examined, indicating that disease may commonly be caused by simultaneous infection with multiple strains. It was concluded that there are a number of different strains of R. equi which carry the vapA gene, and these strains tend to be widespread, but individual farms tend to have particular strains associated with them.

16S rDNA sequence and pyrolysis mass spectrometric analyses were carried out on representatives of Rhodococcus equi and marker strains of genera that encompass mycolic acid containing actinomycetes. The R. equi strains formed a monophyletic clade within the evolutionary radiation occupied by members of the genera Nocardia and Rhodococcus. The 16S rDNA sequence data also showed R. equi to be an heterogeneous taxon. This heterogeneity was underscored by the pyrolysis mass spectrometric data. These observations are in line with those of previous studies where similar profiles of relatedness were found between pyrolysis mass spectral data and the results of DNA:DNA pairing and numerical phenetic studies.

This study was conducted to evaluate the efficacy of the mini-VIDAS Listeria monocytogenes (LMO) system (BioMérieux Vitek, Inc., Missouri, USA) for detection of L. monocytogenes in environmental and fish samples from three Portuguese cold-smoking plants and from their fresh fish suppliers. Mini-VIDAS-LMO is a fully automated system that uses fluorescent ELFA (Enzyme Linked Fluorescent Assay) technology for detection of Listeria monocytogenes antigens in food. It can be a rapid screening method alternative to time consuming classical isolation and identification. Two hundred and ninety five samples were tested in mini-VIDAS-LMO and in parallel by the ISO 11290-1 traditional protocol. The mini-VIDAS-LMO detected 8 of the 11 confirmed positive samples and presented 11 false positive results. The specificity of the mini-VIDAS-LMO found in this experiment was 0.96 and the sensitivity 0.73.

The cultural, morphological, biochemical, serological characteristics and antibiotic susceptibility of 25 Rhodococcus equi strains isolated from lungs and lung abscesses of pneumonic foals and 5 R. equi strains isolated from immuno-compromised human patients were examined. All R. equi strains showed common cultural, morphological and biochemical characteristics both with conventional tests and on the basis of their enzyme profile. The R. equi strains examined were resistant to penicillins with the exception of ampicillin, to sulphamethazine and several strains also to sulphamethoxazole-trimethoprim. All strains were susceptible to erythromycin and rifampicin. The strains isolated from humans showed somewhat higher rate of antibiotic resistance to penicillin, cefotaxime, kanamycin, streptomycin, lincomycin, and oxytetracycline. The overwhelming majority (96%) of the equine isolates belonged to serotype 1 in Prescott's serotyping system, while the human isolates could not be serotyped.

Diversity of virulence-associated antigens of Rhodococcus equi was detected among thirteen strains isolated from AIDS patients on two continents. One out of four Brazilian isolates presented the virulence-associated antigen of 15- to 17-kDa, and the other three isolates had the 20-kDa virulence-associated antigen. In contrast, only three out of nine Italian isolates were positive for virulence-associated antigens - two for the 15- to 17-kDa antigen and one for the 20-kDa antigen. In four other Italian strains, one or more other low-molecular-weight antigens were identified. Because of R. equi variability and host immune dysfunction, no characteristic antibody profile was detected among patients, although the presence of specific antibodies in serum samples suggested prognostic value: good patient outcome and recovery from pneumonia were correlated with R. equi antibody detection, whereas the lack or disappearance of specific antibodies, mainly those to low-molecular-weight antigens, was correlated with disease progression and patient death. These results confirmed the nonobligatory presence of the well-known virulence-associated antigens for the pathogenicity of R. equi in humans, and also the diversity of R. equi strains isolated from AIDS patients, which may be related to the geographic origin of the isolates or may be a consequence of the route of R. equi transmission in different countries. Some mechanisms underlying the results obtained are discussed, suggesting immune complex formation during the progress of the disease.

Pulsed field gel electrophoresis of restriction endonuclease digested genomic DNA from a collection of clinical isolates of Rhodococcus equi was used to compare strain diversity on different Thoroughbred horse farms over time. Restricted diversity was found among the isolates tested, as the same strains were detected on multiple farms and in multiple years. Marked variation occurred in strain prevalence with some strains being represented by single isolates, and the most prevalent by 26 isolates. There were dominant strains on some farms and the prevalence of some strains differed between farms. Infection with multiple strains was noted in some cases where multiple isolates from a single foal were examined.

The present study was designed to comparatively investigate 21 Rhodococcus equi isolates from the faeces of clinically healthy horses and cattle. The isolates were identified by cultural and biochemical properties and by PCR analysis. The latter, targeted to the gene coding for the 16S ribosomal RNA, revealed a species specific PCR product. The isolates were further characterised by serotyping with two typing systems, by haemagglutination tests and by plasmid and virulence protein profiling. Among the 21 cultures, four cultures contained plasmids, two of the four cultures expressed 15-17 kDa virulence proteins, no cultures contained 20 kDA virulence proteins. The 21 cultures were further analysed by DNA-fingerprinting. This was performed by macrorestriction analysis of chromosomal DNA by pulsed-field gel electrophoresis (PFGE). The DNA-restriction patterns were different for most of the isolates indicating a clone heterogeneity among isolates from single farms. Serotyping, determination of virulence marker and PFGE analysis of R equi appeared to be useful for further characterisation of this species, possibly of importance for virulence estimation of single R equi isolates and for epidemiological studies.

Twelve foals, between 27 and 83 days old, were infected with 2 strains of Rhodococcus equi by intratracheal administration. Ten of the 12 foals were inoculated with 10(4)-10(10) colony forming units (cfu) of ATCC 33701 strain. The other 2 foals were inoculated with 10(9) cfu of a plasmid-cured derivative of the ATCC 33701 strain (ATCC 33701P-). All of the 10 foals challenged with the ATCC 33701 strain showed clinical signs of pulmonary disease within 5-13 days, such as gross lesions associated with acute bronchopneumonia and microscopic lesions associated with granulomatous pneumonia. The two foals challenged with the ATCC 33701P- strain showed neither clinical signs of disease nor gross lesions. Apparently, when lacking plasmid, the virulent Rhodococcus equi lost its pathogenicity.

Two different, inactivated, aluminium salt adsorbed vaccines, one containing a R. equi strain (serotype 1, 10(9) CFU/ml and equine herpesvirus 2 (EHV-2) (1.5 x 10(7) PFU/ml) and another containing R. equi only were used on three studfarms to determine whether the disease can be prevented by vaccination of both pregnant mares and their foals. Pregnant mares received two 3 ml doses of vaccine intramuscularly 6 and 2 weeks before parturition and their foals were vaccinated on two or three occasions at 3, 5 or 7 weeks of age. The efficacy of the vaccines was evaluated on the basis of the clinical signs, serological response (indirect haemagglutination and virus neutralisation tests) and culture of R. equi from sick or dead foals. On studs A and B where the bivalent vaccine was used, 24 and 14 foals were born respectively to the vaccinated mares but no clinical case or death occurred due to R. equi pneumonia, while out of the 10 nonvaccinated control foals (stud B) two succumbed to R. equi pneumonia and 4 other foals had to be treated with antibiotics because of fever, coughing and dyspnea. In stud C, where the vaccine containing R. equi strain alone was used, all 15 vaccinated foals remained healthy but one of the 11 control foals died of suppurative R. equi pneumonia and one foal had to be treated due to R. equi pneumonia. R. equi strains (serotype 1) were isolated from the lungs of all dead foals. The serological response was very weak to both R. equi and the EHV-2 strain. Antibody titres in the colostrum of the vaccinated mares against R. equi (in studs A and B, geometric mean 3.79 +/- 1.63 and 4.14 +/- 1.46, respectively) were practically not higher than titres in the controls (in stud B geometric mean 2.12 +/- 1.96). More antibody was present in the colostrum samples against EHV-2 (geometric mean 6.1 + 1.4 compared to 2.5 +/- 1.2). In all foals antibody levels were hardly detectable against both R. equi and EHV-2 until five weeks of age. From the fifth week, antibody levels gradually increased and by the ninth week their reached a titre of 5.5 +/- 1.8 (2.7 +/- 1.2 in the control foals) against R. equi and 5.2 +/- 1.4 against EHV-2. The favorable clinical results and the low antibody titres in the sera of the vaccinated foals during the first week of life suggest that protection probably was due to repeated vaccination of young foals rather than to vaccination of mares.

Rhodococcus equi and Arcanobacterium haemolyticum, formerly classified in the genus Corynebacterium, are members of the loosely defined taxon "coryneform" bacteria. Although they are the etiologic agents of distinct human infections, both organisms are frequently overlooked, which results in missed or delayed diagnoses. R. equi, long known as an important pathogen of immature horses, has become in the past three decades an opportunistic pathogen of severely immunosuppressed humans. Most cases are secondary to HIV infection. When specifically sought in throat swab cultures, A. haemolyticum is found responsible for 0.5% to 2.5% of bacterial pharyngitis, especially among adolescents. These two microorganisms represent a spectrum of disease in humans: from a mild, common illness to a rare life-threatening infection. Each organism elaborates lipid hydrolyzing enzymes (cholesterol oxidase by R. equi and sphingomyelinase D by A. haemolyticum) that are toxic to animals and humans and damaging to mammalian cell membranes. The participation of the cytotoxins in pathogenicity is discussed. Greater awareness of the properties of these two bacteria may promote faster, more accurate diagnoses and better clinical management.

In the present investigation 19 and 22 R. equi-cultures isolated from diseased horses and humans, respectively, could be correctly identified by their morphological, biochemical and serological properties. The rod-coccus life cycle appeared to be a common feature of almost all cultures investigated. The cultures were typeable with the typing system described by Prescott (1981). The predominant serotypes among the R. equi-isolates belonged to serotypes 1 and 2. Among the R. equi-isolates from horses haemagglutination-positive cultures were mainly found among isolates of serotype 1, those of serotype 2 were haemagglutination-negative. The R. equi-cultures isolated from humans showed no relation between serotype and haemagglutinating properties. Determination of antibiotic susceptibility revealed that all cultures were susceptible to erythromycin, gentamicin, imipenem, minocycline, neomycin, rifampicin, streptomycin and vancomycin, 71% and 75% were resistant or at least moderate susceptible to tetracycline and penicillin G, respectively. Almost all cultures were resistant to ceftazidime and most cultures were susceptible to cefotaxime. The cultures could be further characterized by restriction endonuclease digestion and pulsed field gel electrophoresis of their chromosomal DNA. After digestion with the restriction enzyme AsnI the resulting DNA-profile allowed a strain-specific characterization.

The discrepancies between the current description of the CAMP test between Listeria monocytogenes and Rhodococcus equi in the latest edition of Bergey's Manual of Determinative Bacteriology (L. monocytogenes is described as CAMP test negative with R. equi) and routine findings (positive reactions are usually described in many laboratories) make it advisable to review the current interpretation of the CAMP test to avoid confusion among people working in microbiological laboratories. Overall, 98.4% of the L. monocytogenes strains examined in this study, regardless of their source or the intensity of their hemolytic activity, displayed a synergic hemolytic reaction (CAMP phenomenon) with R. equi, indicating that L. monocytogenes can generally be considered CAMP positive with R. equi. We propose that L. monocytogenes, together with Listeria ivanovii, should be considered CAMP test positive with R. equi (circular or racket and semicircular or shovel shapes, respectively).

Rhodococcus equi is an emerging opportunistic pathogen of human immunodeficiency virus-infected patients. However, little is known about the distribution of virulent and intermediately virulent R. equi in human environment. In the present study, R. equi was isolated from 173 of 234 (73.9%) samples collected from soil and sand on 115 parks and 49 yards in Japan. The numbers of R. equi from soil and sand ranged from 2.5 x 10(1) to 1.2 x 10(5) per gram of sample. None of 1,294 isolates from those samples showed virulence-associated 15- to 17-kDa antigens and a 20-kDa antigen. These results suggest that avirulent R. equi is widespread in parks and yards, but the human environment has not been contaminated with virulent and intermediately virulent R. equi strains yet.

Rhodococcus equi is emerging as a cause of pneumonia in immunocompromised people, especially those with AIDS. Like mycobacteria, R. equi is phagocytosed by alveolar macrophages and replicates within them. Recent work is beginning to elucidate the cell and molecular biology of this opportunistic pathogen and the host immune response to it.

In the present study, 17 Rhodococcus equi isolates from animals and 13 R. equi isolates from humans were correctly identified by the use of the API Coryne-test system. The biochemical characteristics revealed no significant differences between isolates from animals and humans. Heat extractions at acid pH or autoclave extractions of the bacteria and the use of monospecific antisera against type antigens 1-7 allowed the serotyping of the bacteria. The majority of the R. equi isolates reacted with serotype 1 and serotype 2-specific antisera, extracts of two human isolates reacted with serotype 5 and serotype 6-specific antisera, respectively. The determination of biochemical and serological properties of R. equi might help to characterize individual cultures of this species. This might be useful in epidemiological studies.