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      Development of a real-time SYBR Green PCR assay for the rapid detection of Dermatophilus congolensis

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          Abstract

          Methods such as real time (RT)-PCR have not been developed for the rapid detection and diagnosis of Dermatophilus ( D.) congolensis infection. In the present study, a D. congolensis-specific SYBR Green RT-PCR assay was evaluated. The detection limit of the RT-PCR assay was 1 pg of DNA per PCR reaction. No cross-reaction with nucleic acids extracted from Pseudomonas aeruginosa, Mycobacterium tuberculosis, Staphylococcus aureus, or Austwickia chelonae was observed. Finally, the RT-PCR assay was used to evaluate clinical samples collected from naturally infected animals with D. congolensis. The results showed that this assay is a fast and reliable method for diagnosing dermatophilosis.

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          Clinical virology in real time.

          Hubert Niesters (2002)
          The ability to detect nucleic acids has had and still has a major impact on diagnostics in clinical virology. Both quantitative and qualitative techniques, whether signal or target amplification based systems, are currently used routinely in most if not all virology laboratories. Technological improvements, from automated sample isolation to real time amplification technology, have given the ability to develop and introduce systems for most viruses of clinical interest, and to obtain clinical relevant information needed for optimal antiviral treatment options. Both polymerase chain reaction (PCR) and nucleic acid sequence-based amplification (NASBA) can currently be used together with real time detection to generate results in a short turn-around time and to determine whether variants relevant for antiviral resistance are present. These new technologies enable the introduction of an individual patient disease management concept. Within our clinical setting, we have introduced this e.g. for quantitative detection of Epstein-Barr Virus (EBV) in T-dell depleted allogeneic stem cell transplant patients. This enabled us to develop models for pre-emptive anti B-cell immunotherapy for EBV reactivation, thereby effectively reducing not the incidence of EBV-lymphoproliferative disease but the virus related mortality. Furthermore, additional clinically relevant viruses can now easily be detected simultaneously. It also becomes more feasible to introduce molecular testing for those viruses that can easily be detected using classical virological methods, like culture techniques or antigen detection. Prospective studies are needed to evaluate the clinical importance of the additional positive samples detected. It should however be made clear that a complete exchange of technologies is unlikely to occur, and that some complementary technologies should stay operational enabling the discovery of new viruses. The implementation of these molecular diagnostic technologies furthermore warrants the use and introduction of standardized materials as well as participation in international quality control programs. Finally, the use of an internal control throughout the whole procedure not only ensures the accuracy of the results generated, but also is necessary to enable precise quantification of these results and to determine detection thresholds more accurately. Since so many targets do have clinical implications, laboratories might prefer to use universal internal controls before the in-house developed assays should be introduced in clinical virology.
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            Dermatophilus congolensis infection (Dermatophilosis) in animals and man! An update.

            L Zaria (1993)
            Dermatophilus congolensis infection (dermatophilosis) is an acute, subacute or chronic skin disease affecting a wide range of species of animals and man. It is world widely distributed but more prevalent in the humid, tropics and subtropics. The disease is now being reviewed in detail. In the present review, the geographical distribution, history and nomenclature, animal hosts affected, aetiological agent and clinical signs of the disease are discussed extensively. Emphasis is also given on the predisposing factors such as rainfall, humidity, ectoparasites etc. under epizootiology. Pathogenesis, pathology, immunology treatment, control and prophylaxis are other areas well covered. The economic importance of the disease is also stressed and a new approach (biological approach) to treatment and control of the disease is being described in this review. It was concluded that in view of the importance of the disease for the increase of livestock and leather production in the tropical and subtropical region especially in Africa, an international centre for dermatophilosis research is highly needed.
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              Mobilicoccus pelagius gen. nov., sp. nov. and Piscicoccus intestinalis gen. nov., sp. nov., two new members of the family Dermatophilaceae, and reclassification of Dermatophilus chelonae (Masters et al. 1995) as Austwickia chelonae gen. nov., comb. nov.

              T Iwami, Moriyuki Hamada, Takao Iino … (2009)
              Two Gram-positive bacteria, designated strains Aji5-31(T) and Ngc37-23(T), were isolated from the intestinal tracts of fishes. 16S rRNA gene sequence analysis indicated that both strains were related to the members of the family Dermatophilaceae, with 95.6-96.9% 16S rRNA gene sequence similarities. The family Dermatophilaceae contains 2 genera and 3 species: Dermatophilus congolensis, Dermatophilus chelonae and Kineosphaera limosa. However, it has been suggested that the taxonomic position of D. chelonae should be reinvestigated using a polyphasic approach, because the chemotaxonomic characteristics are not known (Stackebrandt, 2006; Stackebrandt and Schumann, 2000). Our present study revealed that strains Aji5-31(T), Ngc37-23(T) and D. chelonae NBRC 105200(T) should be separated from the other members of the family Dermatophilaceae on the basis of the following characteristics: the predominant menaquinone of strain Aji5-31(T) is MK-8(H(2)), strain Ngc37-23(T) possesses iso- branched fatty acids as major components, and the menaquinone composition of D. chelonae is MK-8(H(4)), MK-8 and MK-8(H(2)) (5 : 3 : 2, respectively). On the basis of these distinctive phenotypic characteristics and phylogenetic analysis results, it is proposed that strains Aji5-31(T) and Ngc37-23(T) be classified as two novel genera and species of the family Dermatophilaceae. The names are Mobilicoccus pelagius gen. nov., sp. nov. and Piscicoccus intestinalis gen. nov., sp. nov., and the type strains are Aji5-31(T) (=NBRC 104925(T) =DSM 22762(T)) and Ngc37-23(T) (=NBRC 104926(T) =DSM 22761(T)), respectively. In addition, D. chelonae should be reassigned to a new genus of the family Dermatophilaceae with the name Austwickia chelonae gen. nov., comb. nov.
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                Author and article information

                Journal
                Journal ID (nlm-ta): J Vet Sci
                Journal ID (iso-abbrev): J. Vet. Sci
                Journal ID (publisher-id): JVS
                Title: Journal of Veterinary Science
                Publisher: The Korean Society of Veterinary Science
                ISSN (Print): 1229-845X
                ISSN (Electronic): 1976-555X
                Publication date (Print): December 2013
                Publication date (Electronic): 19 December 2013
                Volume: 14
                Issue: 4
                Pages: 491-494
                Affiliations
                Departamento de Sanidad Animal, Facultad de Veterinaria de Cáceres, Universidad de Extremadura, 10003 Cáceres, Spain.
                Author notes
                Corresponding author: Tel: +34-927257114; Fax: +34-927257110; alfrgcia@ 123456unex.es
                Article
                DOI: 10.4142/jvs.2013.14.4.491
                PMC ID: 3885745
                PubMed ID: 23820221
                SO-VID: c68b4103-9973-408a-8135-31044ebd9523
                Copyright © © 2013 The Korean Society of Veterinary Science.
                License:

                This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License ( http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                Date received : 07 February 2013
                Date revision received : 04 April 2013
                Date accepted : 28 June 2013
                Categories
                Subject: Short Communication

                ScienceOpen disciplines: Veterinary medicine
                Keywords: clinical samples,dermatophilus congolensis,dermatophilosis,diagnosis,real-time pcr
                Data availability:
                ScienceOpen disciplines: Veterinary medicine
                Keywords: clinical samples, dermatophilus congolensis, dermatophilosis, diagnosis, real-time pcr

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