mydia acanthamoebae is associated with respiratory diseases in humans and ruminants, as well as diseases of the reproductive organs leading to abortions, or the birth of weak or non-viable offspring. Unfortunately, there are no such test systems in the arsenal of humane and veterinary medicine of Ukraine, which could identify and differentiate Сhlamydia-related organisms, Parachlamydia acanthamoebae in particular. In this regard, PCR test system has been developed for the identification and species differentiation of Parachlamydia acanthamoebae, the bacteria of the Parachlamydiaceae family, which is the causative agent of chlamydiosis in ruminants and humans. Conducted bioinformatics studies revealed species-specific regions of the Parachlamydia acanthamoebae genome, which were used to develop the nucleotide structure of oligonucleotide primers, which allow to amplify the DNA fragment of the 16S rRNA gene specific for the bacterium Parachlamydia acanthamoebae, the size of 88 base pairs. Verification of the analytical specificity of the PCR test system was confirmed by amplifying 9 control DNA of members of the order Chlamydiales: Waddlia chrondophila, Clavochlamydia salmonicola, Piscichlamydia salmonis, Chlamydia avium, Chlamydia pecorum, Chlamydia abortus, Chlamydia psittaci, Chlamydia suis, Chlamydia caviae. The demand for such test systems is due not only to their use in scientific research to isolate and study individual strains, to conduct epizootic monitoring, but also for the purpose of effective treatment, since distinct species have diverse sensitivity or even insensitivity to different types of antibiotics and vary in the source of infection and mode of transmission.
Мета статті – надати актуальну на сьогоднішній день таксономічну класифікацію порядку Chlamydiales та детально роз’яснити терміни: «Chlamydia-related bacteria (CRB)» (хламідіє-споріднені бактерії,) «Chlamydia-like organisms (CLO)» (хламідіє-подібні організми), «environmental chlamydiae» (екологічні хламідії, хламідії довкілля). Методика дослідження. Використано такі методи дослідження: системний аналіз доступних наукових джерел, історичний метод (для вивчення виникнення, формування та розвитку таксономії хламідійних видів у хронологічній послідовності), емпіричний метод (щодо комплексної оцінки сучасного стану об’єкта дослідження), абстрактно-логічний ‒ для уточнення суті основних понять та графічне відображення даних. Результати дослідження. Внаслідок розвитку молекулярної біології за останні 15 років класифікація хламідій зазнала істотних змін і має тенденцію до подальшого вдосконалення. У статті узагальнено історичний досвід і найбільш важливі зміни в таксономії порядку Chlamydiales за останні роки. Ускладнення сучасної класифікації бактерій порядку Chlamydiales свідчить про накопичені знання щодо нових представників цієї групи мікроорганізмів. На сьогодні до порядку Chlamydiales входять 9 родин: Chlamydiaceae, Waddliaceae, Parachlamydiaceae, Criblamydiaceae, Simkaniaceae, Candidatus Clavochlamydiaceae, Candidatus Rhabdochlamydiaceae, Candidatus Piscichlamydia, Candidatus Parilichlamydiaceae, чотири з яких перебувають у статусі кандидатів. До родини Chlamydiaceae, роду Chlamydia тепер належать 14 видів бактерій (Chlamydia abortus, Chlamydia avium, Chlamydia caviae, Chlamydia felis, Chlamydia gallinacea, Chlamydia muridarum, Chlamydia pecorum, Chlamydia pneumoniae, Chlamydia psittaci, Chlamydia suis, Chlamydia trachomatis, Candidatus Chlamydia ibidis, Candidatus Chlamydia sanzinia, Candidatus Chlamydia corallus), три з яких перебувають у статусі кандидатів. Родини: Waddliaceae, Parachlamydiaceae, Criblamydiaceae, Simkaniaceae, Ca. Clavochlamydiaceae, Ca. Rhabdochlamydiaceae, Ca. Piscichlamydia, Ca. Parilichlamydiaceae з їхніми численними представниками називають хламідіє-спорідненими бактеріями, або хламідіє-подібними організмами, через їх генетичну і фенотипову подібність та філогенетичну відокремленість від родини Chlamydiaceae. Елементи наукової новизни. У цій статті представлено актуальну таксономічну класифікацію порядку Chlamydiales та надано детально роз’яснено терміни: «Chlamydia-related bacteria (CRB)» (хламідіє-спорідненені бактерії,) «Chlamydia-like organisms (CLO)» (хламідіє-подібні організми), «environmental chlamydiae» (екологічні хламідії, хламідії довкілля). Практична значущість. Надана інформація може бути використана в наукових дослідженнях, впроваджена в навчальний процес при підготовці спеціалістів у галузі ветеринарної медицини, а також використовуватися практикуючими лікарями ветеринарної медицини задля покращення лікування та профілактики хламідіозів тварин та людини. The purpose of the article is to provide the currently relevant taxonomic classification of the Order Chlamydiales, and give a detailed explanation of the terms: “Chlamydia-related bacteria (CRB)”, “Chlamydia-like organisms (CLO)”, "Environmental chlamydiae" (ecological chlamidiae). Research methods. The following research methods were used: system analysis of the available scientific sources, historical method (to study the origin, formation and development of the taxonomy of chlamydia species in chronological order), empirical method (concerning the comlex assessment of the current state of the research object), abstract-logical – to clarify the essence of the basic notions and graphical reflection of the data. The results of the study. As a result of molecular biology development during the recent 15 years, the classification of chlamydia has undergone significant changes and has the tendency to further improvement. This review article is aimed at summarizing the historical experience and the most important changes in the taxonomy of the Order Chlamydiales in recent years. The complication of the modern classification of the Order Chlamydiales bacteria is the inevitable consequence of accumulating knowledge about the new representatives of this group of microorganisms. Consequently at present there are 9 families in the Order Chlamydiales: Chlamydiaceae, Waddliaceae, Parachlamydiaceae, Criblamydiaceae, Simkaniaceae, Candidatus Clavochlamydiaceae, Candidatus Rhabdochlamydiaceae, Candidatus Piscichlamydia, Candidatus Parilichlamydiaceae, four of them are in the status of candidates. Currently, 14 species of bacteria belong to the family Chlamydiaceae, the genus Chlamydia (Chlamydia abortus, Chlamydia avium, Chlamydia caviae, Chlamydia felis, Chlamydia gallinacea, Chlamydia muridarum, Chlamydia pecorum, Chlamydia pneumoniae, Chlamydia psittaci, Chlamydia suis, Chlamydia trachomatis, Candidatus Chlamydia ibidis, Candidatus Chlamydia sanzinia, Candidatus Chlamydia corallus), three of which are the candidates. Such families as: Waddliaceae, Parachlamydiaceae, Criblamydiaceae, Simkaniaceae, Ca. Clavochlamydiaceae, Ca. Rhabdochlamydiaceae, Ca. Piscichlamydia, Ca. Parilichlamydiaceae with their numerous representatives are called Сhlamydia-related bacteria, or Сhlamydia-like organisms, due to their genetic and phenotypic similarity, and phylogenetic isolation from the Chlamydiaceae family. The elements of scientific novelty. The currently relevant taxonomic classification of Chlamydiales is presented in the article and also given a detailed explanation of the terms: “Chlamydia-related bacteria (CRB)”, “Chlamydia-like organisms (CLO)”, "Environmental chlamydiae". Practical significance. The provided information can be used in scientific research, introduced into the educational process for training specialists of veterinary medicine, and also be used by practicing veterinary doctors in order to improve the treatment and prevention of animals and human chlamydioses.
The aim of our work was to survey private farms in the Poltava region on the prevalence of Chlamydia-like organisms Waddlia chondrophila and Parachlamydia acanthamoebae. Materials and methods: The PCR research was conducted in 2018-2019 at the Institute of Pig Breeding and Аgro-industrial production of the NAAS. The study materials were 300 vaginal scrapings of cows and 28 samples of clinical material of pigs and boars with a suspicion of a chlamydial infection, which were selected at private farms in the Poltava region. Results: Chlamydia-like bacteria Waddlia chrondophila DNA was detected in 132 (44%) samples taken from cows and 8 (28.6%) samples from pigs and boars. Parachlamydia acanthamoebae DNA was not detected in pig samples. However, 43 (14.3%) clinical samples taken from cows were found to be Parachlamydia acanthamoebae – positive. 9 (3%) cows and 2 (7.14%) pigs turned out to be Chlamydia spp. – positive (the samples contained DNA of bacteria of the family Chlamydiaceae). Сonclusions: Our data indicate a relatively high prevalence of Chlamydia-like bacteria in problematic as to chlamydial infection farms in the Poltava region. Chlamydial infection carry a zoonotic threat, therefore it is necessary to observe safety and personal hygiene measures when working with animals. In addition, it is necessary to carry out periodic screening among the population with reproductive disorders and in occupational risk groups, periodic monitoring in livestock farms, to avoid the consuming the unpasteurized milk, and thermally unprocessed meat.
Epitheliocystis is an emerging disease of wild and cultured fish caused by a number of bacterial species, characterized by the presence of cytoplasmic bacterial inclusions in the epithelial cells of the gills, which contribute to the merging of the gill plates, and in some cases also spread to the skin of fish. This disease may manifest as hypertrophy and inflammation of the gills, white nodular lesions of epithelial tissue in the gills or skin, gasping on the surface of the water, lethargy, poor swimming and stunted growth. Among the commercially important aquaculture species of Ukraine, such as Atlantic salmon (Salmo salar), brown trout (S. trutta), grass carp (Ctenopharyngodon idella), common carp (Cyprinus carpio) and gibel carp (Carassius auratus), Candidatus Clavochlamydia salmonicola and Candidatus Piscichlamydia salmonis are associated with epitheliocystis. There are currently no tools at the disposal of ichthyologists and veterinary laboratories in Ukraine to identify Ca. C. salmonicola and Ca. Piscichlamydia salmonis. Our basic concern was to develop a PCR assay of epitheliocystis diagnosis. We suggest the use of general primers for simultaneous detection of Ca. C. salmonicola and Ca. Piscichlamydia salmonis. The developed PCR assay for identification of Ca. C. salmonicola and Ca. Piscichlamydia salmonis has shown its suitability for amplifying control DNA. Confirmation of the amplification products identity was performed using selective recognition of the sequence by the TasI restriction endonuclease (Thermo Fisher Scientific, US). Analytical specificity verification of the PCR assay performed by amplifying the control DNA of 10 species of the Chlamydiales order showed the absence of PCR products, but observed in one. The designed PCR assay, after approbation on clinical material, can be used by researchers for extensive monitoring of epitheliocystis, doctors of veterinary medicine for diagnosis clarification, in addition to introduction into the practice of veterinary medicine laboratories and implementation in fish farm improvement programmes. The amplicon size of 197 base pairs theoretically permits application of this oligonucleotide primers pair for real-time PCR.
The aim of our work was to develop PCR test systems for the identification and differentiation of the Piscichlamydia salmonis and Clavochlamydia salmonicola, species, that are known epitheliocystis infection agents of gill and fish skin diseases, characterized by the presence of specific ‘inclusions’ in the epithelial cells of the gills. To date, the diseases of fish associated with chlamydial infections have been detected in more than 90 species of freshwater and marine fish worldwide. For now, there is no available information on the prevalence of Piscichlamydia salmonis and Clavochlamydia salmonicola, which can cause epitheliocystis of commercially important aquaculture species in Ukraine. Identification of these pathogens is possible only using molecular genetic methods. As a result of our research, we got PCR tests for the identification and species differentiation of Piscichlamydia salmonis and Clavochlamydia salmonicola. The use of diagnostics for the identification of Piscichlamydia salmonis and Clavochlamydia salmonicola makes chlamydial infections monitoring among various fish species possible and it will increase the economic efficiency of fish farms.
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