Signature-tagged mutagenesis of Vibrio vulnificus

Vibrio vulnificus is the causative agent of primary septicemia, wound infection and gastroenteritis in immunocompromised people. In this study, signature-tagged mutagenesis (STM) was applied to identify the virulence genes of V. vulnificus. Using STM, 6,480 mutants in total were constructed and divided into 81 sets (INPUT pools); each mutant in a set was assigned a different tag. Each INPUT pool was intraperitoneally injected into iron-overloaded mice, and in vivo surviving mutants were collected from blood samples from the heart (OUTPUT pools). From the genomic DNA of mixed INPUT or OUTPUT pools, digoxigenin-labeled DNA probes against the tagged region were prepared and used for dot hybridization. Thirty tentatively attenuated mutants, which were hybridized clearly with INPUT probes but barely with OUTPUT probes, were negatively selected. Lethal doses of 11 of the 30 mutants were reduced to more than 1/100; of these, the lethal doses of 2 were reduced to as low as 1/100,000. Transposon-inserted genes in the 11 attenuated mutants were those for IMP dehydrogenase, UDP-N-acetylglucosamine-2-epimerase, aspartokinase, phosphoribosylformylglycinamidine cyclo-ligase, malate Na (+) symporter and hypothetical protein. When mice were immunized with an attenuated mutant strain into which IMP dehydrogenase had been inserted with a transposon, they were protected against V. vulnificus infection. In this study, we demonstrated that the STM method can be used to search for the virulence genes of V. vulnificus.     

Bacteriological and Molecular Detection of Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus in Equines of Northern India

Present study was undertaken to study the prevalence of β-haemolytic streptococci in equine of northern temperate region of Jammu and Kashmir, India. One hundred and forty one samples were collected in duplicate from nasopharyngeal tract of diseased (53) and apparently healthy equine (88) for isolation and direct PCR. A total of 77 isolates of streptococci were recovered from 141 samples with an overall prevalence rate of 54.60%. Out of these 77 isolates, 52 were from diseased and 25 from apparently healthy animals. Of the 77 isolates, 4 were identified as Streptococcus equi subsp. equi, 56 as S. equi subsp. zooepidemicus and 17 as S. dysgalactiae subsp. equisimilis. Thus the overall prevalence of S. equi subsp. equi, S. equi subsp. zooepidemicus and S. dysgalactiae subsp. equisimilis was 2.83, 39.71 and 12.05% respectively. The sensitivity of the PCR for the detection of S. equi species was found higher when attempted from direct swab samples.     

Anti-emetic drug maropitant induces intestinal motility disorder but not anti-inflammatory action in mice

Maropitant is a neurokinin 1 receptor (NK1R) antagonist that is clinically used as a new anti-emetic drug for dogs. Substance P (SP) and its receptor NK1R are considered to modulate gastrointestinal peristalsis. In addition, SP works as an inflammatory mediator in gastrointestinal diseases. Aim of this study is to clarify the effects of maropitant on intestinal motility and inflammation in mice. Ex vivo examination of luminal pressure-induced intestinal motility of whole intestine revealed that maropitant (0.1–10 µM) increased frequency of contraction, decreased amplitude of contraction and totally inhibited motility index in a concentration-dependent manner. We measured intestinal transit in vivo by measuring transportation of orally administered luminal content labeled with phenol red. Our results demonstrated that maropitant (10 mg/kg, SC) delayed intestinal transit. Geometric center value was significantly decreased in maropitant-treated mice. Anti-inflammatory effects of maropitant against leukocytes infiltration into the intestinal smooth muscle layer in post-operative ileus (POI) model mice were measured by immunohistochemistry. In POI model mice, a great number of CD68-positive macrophages or MPO-stained neutrophils infiltrated into the inflamed muscle region of the intestine. However, in the maropitant treated mice, the infiltration of leukocytes was not inhibited. The results indicated that maropitant has ability to induce disorder of intestinal motility in mice, but has no anti-inflammatory action in the mouse of a POI model. In conclusion, in mice, maropitant induces disorder of intestinal motility in vivo.     

Lineages of Streptococcus equi ssp. equi in the Irish equine industry

Background

Streptococcus equi ssp. equi is the causative agent of ‘Strangles’ in horses. This is a debilitating condition leading to economic loss, yard closures and cancellation of equestrian events. There are multiple genotypes of S. equi ssp. equi which can cause disease, but to date there has been no systematic study of strains which are prevalent in Ireland. This study identified and classified Streptococcus equi ssp. equi strains isolated from within the Irish equine industry.

Results

Two hundred veterinary isolates were subjected to SLST (single locus sequence typing) based on an internal sequence from the seM gene of Streptococcus equi ssp equi. Of the 171 samples which successfully gave an amplicon, 162 samples (137 Irish and 24 UK strains) gave robust DNA sequence information. Analysis of the sequences allowed division of the isolates into 19 groups, 13 of which contain at least 2 isolates and 6 groups containing single isolates. There were 19 positions where a DNA SNP (single nucleotide polymorphism) occurs, and one 3 bp insertion. All groups had multiple (2–8) SNPs. Of the SNPs 17 would result in an amino acid change in the encoded protein. Interestingly, the single isolate EI8, which has 6 SNPs, has the three base pair insertion which is not seen in any other isolate, this would result in the insertion of an Ile residue at position 62 in that protein sequence. Comparison of the relevant region in the determined sequences with the UK Streptococcus equi seM MLST database showed that Group B (15 isolates) and Group I (2 isolates), as well as the individual isolates EI3 and EI8, are unique to Ireland, and some groups are most likely of UK origin (Groups F and M), but many more probably passed back and forth between the two countries.

Conclusions

The strains occurring in Ireland are not clonal and there is a considerable degree of sequence variation seen in the seM gene. There are two major clades causing infection in Ireland and these strains are also common in the UK.     

Transposon mutagenesis in Mycoplasma hyopneumoniae using a novel mariner-based system for generating random mutations

Mycoplasma hyopneumoniae is the cause of enzootic pneumonia in pigs, a chronic respiratory disease associated with significant economic losses to swine producers worldwide. The molecular pathogenesis of infection is poorly understood due to the lack of genetic tools to allow manipulation of the organism and more generally for the Mycoplasma genus. The objective of this study was to develop a system for generating random transposon insertion mutants in M. hyopneumoniae that could prove a powerful tool in enabling the pathogenesis of infection to be unraveled. A novel delivery vector was constructed containing a hyperactive C9 mutant of the Himar1 transposase along with a mini transposon containing the tetracycline resistance cassette, tetM. M. hyopneumoniae strain 232 was electroporated with the construct and tetM-expressing transformants selected on agar containing tetracycline. Individual transformants contained single transposon insertions that were stable upon serial passages in broth medium. The insertion sites of 44 individual transformants were determined and confirmed disruption of several M. hyopneumoniae genes. A large pool of over 10 000 mutants was generated that should allow saturation of the M. hyopneumoniae strain 232 genome. This is the first time that transposon mutagenesis has been demonstrated in this important pathogen and could be generally applied for other Mycoplasma species that are intractable to genetic manipulation. The ability to generate random mutant libraries is a powerful tool in the further study of the pathogenesis of this important swine pathogen.     

Changes in Serum Antibody Levels after Vaccination for Strangles and after Intranasal Challenge with Streptococcus equi subsp. equi in Horses

In this study, to evaluate the influence of strangles vaccination on serological test results, we investigated the changes in strangles serum antibody levels in horses after vaccination and subsequent intranasal challenge with S. equi. The horses were vaccinated for strangles with either a component vaccine (Group C) or a live vaccine (Group L). We measured changes in strangles serum antibody levels weekly for 20 weeks after vaccinating horses twice for strangles over a 3-week interval, and for 7 weeks after intranasal challenge with S. equi in the same horses. Serum antibody responses to the proline-glutamic acid-proline-lysine (PEPK) antigen with five repetitions (PEPK-5R) were higher at all times (up to 2.4-fold) following vaccination in Group C than in Group L, and the value peaked at 2.9-fold above the initial value after the second vaccination in Group C horses. However, the value was lower than that in horses infected with S. equi, and it gradually decreased, reaching the initial (week 0) value by the 15th week. Serum antibody responses to PEPK-5R after challenge with S. equi increased in both groups of horses, but the value tended to be lower than that reported for unvaccinated horses. In addition, the average value in Group C was 2.6-fold higher than that of Group L. These results suggest the serum antibody responses of horses infected with S. equi varies according to the type of vaccine with which they have been vaccinated. Although the serological diagnostic test for strangles in which PEPK-5R is used as an antigen is effective for the investigation of serum antibodies to strangles in vaccinated horses, the present data suggest it is necessary to consider the vaccination history when interpreting the results.     

Rhodococcus equi (Prescottella equi) vaccines; the future of vaccine development

For decades researchers have been targeting prevention of Rhodococcus equi (Rhodococcus hoagui/Prescottella equi) by vaccination and the horse breeding industry has supported the ongoing efforts by researchers to develop a safe and cost effective vaccine to prevent disease in foals. Traditional vaccines including live, killed and attenuated (physical and chemical) vaccines have proved to be ineffective and more modern molecular‐based vaccines including the DNA plasmid, genetically attenuated and subunit vaccines have provided inadequate protection of foals. Newer, bacterial vector vaccines have recently shown promise for R. equi in the mouse model. This article describes the findings of key research in R. equi vaccine development and looks at alternative methods that may potentially be utilised.     

Prevalence and Significance of Parasites of Horses in Some States of Northern Nigeria

This study was conducted to determine the prevalence and significance of parasites of horses in northern Nigeria. Blood and faecal samples were randomly collected from 243 horses from different stables in some states of northern Nigeria for laboratory analyses. Fifty-seven horses (23.5%) were found infected with parasites. The hemoparasites detected, 21 (8.6%), include Theileria equi, Babesia caballi, Trypanosoma vivax and Trypanosoma evansi. The endoparasites encountered, 29 (11.9%) were Strongylus spp., Strongyloides spp., Oxyuris equi, Parascaris equorum, Paragonimus spp. and Dicrocoelium spp., 3 (1.2%) was Eimeria spp. Four horses (1.6%) had mixed infection of hemo- and endoparasites. This preliminary finding shows that parasitism is a problem in the horse stables examined, and calls for proper stable hygiene, routine tick control and regular deworming programme.     

Serological epidemiological surveillance for vapN-harboring Rhodococcus equi infection in goats

Rhodococcus equi causes suppurative pneumonia in foals aged 1–3 months; moreover, it has emerged as a pathogenic cause of zoonotic diseases. After the initial report of the ruminant-pathogenic factor VapN encoded by the novel virulence plasmid pVAPN, several reports have described ruminant infections caused by vapN-harboring R. equi. Herein, we conducted a serological epidemiological surveillance in goats at a breeding farm (Farm A) and characterized the vapN-harboring R. equi isolates from this farm. First, we established a simple screening enzyme-linked immunosorbent assay (ELISA) using recombinant glutathione S-transferase–tagged VapN as an immobilized antigen. This method revealed that the VapN antibody titers were elevated in 12 of 42 goats. Subsequently, we attempted to isolate R. equi from the goat feces and soil of Farm A. choE⁺/vapN⁺ R. equi was isolated from the feces of Goat No. 27 and a soil sample near the shed. The pulsed-field gel electrophoresis (PFGE) patterns of five vapN-harboring R. equi strains isolated from Farm A in 2013 and 2019 were investigated and found to be the same except for the strain (OKI2019F1). However, no difference was observed in VapN expression and growth in macrophages among these vapN-harboring R. equi isolates. Our results revealed that some goats had histories of vapN-harboring R. equi infections, and two genomic types of vapN-harboring R. equi were found in isolates from Farm A. Ruminant-specific (pVAPN-carrying) R. equi might be an unrecognized pathogen in Japan and further studies are required to determine its prevalence and distribution.     

Cytokine expression by neutrophils of adult horses stimulated with virulent and avirulent Rhodococcus equi in vitro

Rhodococcus equi is an intracellular pathogen of macrophages that causes rhodococcal pneumonia in foals and immunocompromised people. Evidence exists that neutrophils play a vital role in resistance to infection with R. equi; however, the means by which neutrophils exert their effects have not been clearly defined. In addition to directly killing bacteria, neutrophils also may exert a protective effect by linking innate and adaptive immune responses. In the present study we evaluated the cytokine expression profiles of adult equine neutrophils in response to stimulation with isogenic strains of virulent and avirulent R. equi in vitro. After 2 and 4 h incubation with virulent or avirulent R. equi, adult equine neutrophils expressed significantly (P < 0.05) greater tumor necrosis factor alpha (TNFα), interleukin (IL)-12p40, IL-6, IL-8 and IL-23p19 mRNA, but not interferon gamma (IFNγ) or IL-12p35 mRNA than unstimulated neutrophils. Furthermore, virulent R. equi induced significantly greater IL-23p19 mRNA than avirulent R. equi. These results demonstrate that R. equi-stimulated neutrophils are a source of many proinflammatory cytokines. Furthermore, these results suggest that IL-23 may be preferentially expressed over IL-12 in response to exposure with R. equi, and that this response may be more strongly induced by virulent R. equi than avirulent R. equi. Collectively, the data presented herein suggest a non-phagocytic role for neutrophils that may influence the type of adaptive immune response to R. equi.     

Characterization of a Streptococcus equi ssp. equi Isolate From a Strangles Outbreak in Thailand

Strangles, which is caused by Streptococcus equi ssp. equi, is one of the major infectious respiratory diseases in horses. Knowledge of isolates from different areas of the world is important for investigating the different strains of the disease. In contrast to many other countries, currently little is known about S. equi ssp. equi isolates in Thailand. In 2014, a farm in Thailand imported 20 horses from Europe. Approximately 1 month after arrival, 50% of the horses had developed pyrexia, mucopurulent nasal discharge, and abscesses of the mandibular lymph nodes. Nasal swabs of mucopurulent discharge were sent to a diagnostic laboratory, and two isolates of S. equi ssp. equi were identified. One of the isolates was further characterized using seM gene polymerase chain reaction and sequence analysis. The seM sequence was then compared to the database of PubMLST-seM. It was found to contain SeM allele 48, an allele isolated from horses in the United Kingdom in 2006 and 2010. This result demonstrates the usefulness of SeM allele identification as a tool for investigating the source of related strains and for the epidemiologic study of strangles. To the best of the authors' knowledge, this is the first report of the identification of an SeM allele of the S. equi ssp. equi isolate in Thailand.     

Diagnostic testing patterns for Streptococcus equi subsp. equi in Ontario horses during the years 2008 to 2018

This retrospective study describes testing patterns and the incidence of Streptococcus equi subsp. equi in Ontario to assess the utility of laboratory data for surveillance purposes. Laboratory records for equine infectious disease test submissions were extracted from the Animal Health Laboratory (AHL) at the University of Guelph for the years 2008 to 2018. Yearly and seasonal trends in S. equi testing and the proportion of tests that returned positive results were assessed. The number of samples submitted for S. equi testing decreased over the 11-year period (odds ratio = 0.96, 95% confidence interval: 0.92 to 0.999; P = 0.04). A generalized linear model identified a significant seasonal effect for animals recognized as clinically ill, with the highest test positivity noted in the winter. Although this study identified important trends in the incidence of S. equi in Ontario, the variability in information accompanying test submissions made the data challenging to interpret, highlighting the need for more complete diagnostic submission data for S. equi.     

Assessment of Antimicrobial Susceptibility of Virulent Strains of Rhodococcus equi Isolated From Foals and Soil of Horse Breeding Farms With and Without Endemic Infections

Rhodococcus equi is an opportunistic, intracellular saprophyte that causes severe pyogranulomatous pneumonia in foals. The bacterium displays in vitro susceptibility to many antibiotics. The highest efficacy against R. equi in vitro and in vivo is achieved by using a combination of rifampicin and macrolide antibiotics. Recent years have seen an upward trend in the minimum inhibitory concentration (MIC) of rifampicin and erythromycin, suggesting increasing resistance of R. equi to these antibiotics. The aim of the study was to determine the antimicrobial activity of 24 selected antibiotics against R. equi strains isolated from dead foals and from the environment of horse breeding farms with and without endemic R. equi infections. Minimum inhibitory concentration gradient strips were used to determine the lowest concentration of the antibiotic that inhibited the growth of R. equi. Based on normal MIC distribution, an epidemiologic cutoff values (ECOFF) were assessed for particular antibiotics and R. equi strains. The results were compared with ECOFFs for S. aureus, according to the European Committee on Antimicrobial Susceptibility Testing data. The data indicate that the lowest MIC values were obtained for clarithromycin, rifampicin, imipenem, and vancomycin. The majority of R. equi strains can be classified as wild type in relation to the majority of antibiotics. A small percentage of strains presented non-WT (NWT) with the exception of SXT, for which 35% of strains were classified as NWT. The lack of interpretative criteria for R. equi creates a real problem in the assessment of antibiotic sensitivity both for clinical and scientific purposes.     

Mutagenesis of Streptococcus equi and Streptococcus suis by transposon Tn917

Genetic tools for studying streptococci are much less sophisticated than those that are available for many other bacterial genera. In this paper, we describe the development of a transposon mutagenesis system that we have used to mutate two important veterinary streptococci, Streptococcus equi and Streptococcus suis. The system uses a temperature-sensitive suicide vector to deliver Tn917 via electroporation, transposing Tn917 into the chromosomal DNA of the two streptococci. The transposon insertions can be rescued from the streptococcal chromosomes by plasmid rescue and selection in E. coli, with subsequent insertion site analysis by DNA sequencing. Transposition appeared to have occurred in an essentially random fashion when chromosomal DNA of S. suis and S. equi mutants was analysed by Southern blotting. However, when analysis of 60 S. equi mutants was carried out using the S. equi genome sequence database, 60% of transposon insertions had occurred within a 15 kb region of the genome whereas the other insertions appeared to have occurred essentially randomly. This finding suggests that Southern blot analysis for assessing the randomness of transposon libraries may need to be interpreted with caution. However, this observation notwithstanding, the Tn917 based system described in this paper will facilitate the study of S. suis and S. equi.     

Application of humanized mice to toxicology studies: Evaluation of the human relevance of the mode of action for rodent liver tumor formation by activators of the constitutive androstane receptor (CAR)

The constitutive androstane receptor (CAR)-mediated mode of action (MOA) for phenobarbital (PB)-induced rodent liver tumor formation has been established, with increased hepatocyte proliferation, which is a key event in tumor formation. Previous studies have demonstrated that PB and other CAR-activators stimulate proliferation in cultured rodent hepatocytes, but not in cultured human hepatocytes. However, in the genetically humanized CAR and pregnane X receptor (PXR) mouse (hCAR/hPXR mouse, downstream genes are still mouse), PB increased hepatocyte proliferation and tumor production in vivo. In contrast to the hCAR/hPXR mouse, studies with chimeric mice with human hepatocytes (PXB-mouse, both receptor and downstream genes are human) demonstrated that PB did not increase human hepatocyte proliferation in vivo. PB increased hepatocyte proliferation in a chimeric mouse model with rat hepatocytes, indicating that the lack of human hepatocyte proliferation is not due to any functional defect in the chimeric mouse liver environment. Gene expression analysis demonstrated that the downstream genes of CAR/PXR activation were similar in hCAR/hPXR and CD-1 mice, but differed from those observed in chimeric mice with human hepatocytes. These findings strongly support the conclusion that the MOA for CAR-mediated rodent liver tumor formation is qualitatively implausible for humans. Indeed, epidemiological studies have found no causal link between PB and human liver tumors. There are many similarities with respect to hepatic effects and species differences between rodent CAR and peroxisome proliferator-activated receptor α activators. Based on our research, the chimeric mouse with human hepatocytes (PXB-mouse) is reliable for human cancer risk assessment of test chemicals.     

Associations between the Exposure to Airborne Virulent Rhodococcus equi and the Incidence of R equi Pneumonia among Individual Foals

Rhodococcus equi is a significant cause of pneumonia, resulting in disease and sometimes death of foals. It is believed that infection occurs by inhalation of dust contaminated with virulent R equi. Although association between the airborne concentration of virulent R equi and the incidence of foal pneumonia at breeding farms has been documented, studies at the level of individual foals have not been reported. Thus, the objective of this study was to determine whether the magnitude of airborne virulent R equi was significantly associated with risk of R equi pneumonia for individual foals. The concentration of virulent R equi was significantly (P < .001) greater in stalls than paddocks among samples collected from 47 foals at a breeding farm in central Kentucky. The presence of airborne virulent R equi in stalls was significantly (P = .045) more likely at 7 days of age for foals subsequently found to be affected by rhodococcal pneumonia. Additionally, airborne concentrations of virulent R equi in stalls were significantly greater at 7 and 14 days of age than at birth. Presence of the mare and foal at the time of sampling was significantly (P < .001) associated with increased airborne concentrations of virulent R equi in stalls. These findings suggest that environments containing airborne virulent R equi during the first week of life may influence the risk of subsequent disease for a foal.     

Some problems associated with the identification of Corynebacterium equi

One hundred strains of Corynebacterium equi were examined using standard laboratory procedures. Variations in colonial morphology were noted. The ability to reduce nitrate and to produce urease was shown to be valuable for confirmation of identification of C. equi. Production of hydrogen sulphide was a variable characteristic of C. equi. It is concluded that for those situations where the source of the organism gives no clue as to its identify, positive identification of C. equi may be difficult because of variability in colonial appearance and biochemical reactions.     

Frequency of Rhodococcus equi in Feces of Mares in Central Kentucky

Rhodococcus equi (R equi) pneumonia is an important cause of disease and death in foals. Feces from mares can contain R equi, including virulent R equi, and thus may act as a source of the bacteria at horse breeding farms. A previous report documented that every mare at a farm in central Kentucky shed virulent R equi in at least one of four fecal samples collected serially during the periparturient period. The objective of this study was to assess the extent to which this high prevalence of fecal shedding could be replicated at other horse breeding farms. The frequency of detection of R equi and virulent R equi in fecal samples was studied among 131 mares from 24 farms in central Kentucky. The proportions of fecal samples from mares containing R equi and virulent R equi were 95% and 76%, respectively. These findings indicate that R equi and virulent R equi may be isolated with high frequency from feces of mares at breeding farms in central Kentucky, and that mares are a source of virulent R equi for the environment of their foals.     

Pathology of female mice experimentally infected with an in vitro cultured strain of Trypanosoma equiperdum

Dourine, caused by infection with Trypanosoma equiperdum, is one of the trypanosomiasis in equids. The clinical course of dourine is long-term, ranging from 1–2 months to several years. Since the pathogenesis of dourine has not yet been elucidated, experimental studies using mouse infection models are needed. Although mice are not susceptible to most T. equiperdum strains, some strains can infect mice. Even in such strains, infected mice develop rapidly transient parasitemia and die within 2–8 days. Therefore, mice experimentally infected with these T. equiperdum strains are not suitable for mouse infection models to analysis the pathogenesis of dourine. A sequential method of isolating parasites from dourine-affected horses and adapting them to in vitro cultures using soft agarose media was recently developed. Various T. equiperdum strains adapted to in vitro conditions have been established using this technique. We used one of these strains, the T. equiperdum IVM-t2 strain. In the present study, T. equiperdum IVM-t2 strain inoculated mice developed periodic parasitemia during the experimental period of 60 days. Histopathologically, vaginitis and dermatitis were observed. These findings were comparable to those of dourine-affected horses. Therefore, mice infected with T. equiperdum IVM-t2 strain may be a valuable tool for pathological, immunological, and parasitological in vivo research, and will contribute to investigations on the mechanisms underlying the disease process and the host-parasite relationship.     

Estimating the Sensitivity and Specificity of Real‐Time Quantitative PCR of Fecal Samples for Diagnosis of Rhodococcus equi Pneumonia in Foals

Background

Real‐time, quantitative PCR (qPCR) methods for detecting Rhodococcus equi in feces have been developed as a noninvasive, rapid diagnostic test for R. equi pneumonia, but have not been evaluated in a large population of foals.

Objective

The objective of this study was to evaluate the clinical utility of fecal PCR as a diagnostic test for R. equi pneumonia in foals using receiver operating characteristic (ROC) methods.

Animals

186 foals born in 2011 at an R. equi‐endemic ranch in Texas.

Methods

Fecal samples were collected at the time of onset of clinical signs for pneumonic foals (n = 31). Foals with pneumonia were matched by age and birth date to healthy (n = 31) and subclinical (n = 124) control foals; fecal samples were collected from these controls. DNA was extracted from feces using commercial kits and concentration of virulent R. equi in feces was determined by qPCR.

Results

Concentration of R. equi in feces differed significantly (P < .05) among groups. The area under the ROC curve for fecal qPCR for diagnosis of R. equi pneumonia was 89% (95% CI, 83–99), with a sensitivity of 94% and specificity of 72%.

Conclusions and Clinical Importance

qPCR of feces can be useful as an alternative to tracheobronchial aspiration for the diagnosis of R. equi in foals with clinical signs of pneumonia. Caution should be used in extrapolating results of this study to other populations because fecal concentration of R. equi might vary by geographic location or management practices.