Studyof resistance of pathogenic and opportunistic fungi toantimycotics

The widespread use of antimycotic agents for the treatment of mycoses in humans and animals is of concern to medical and veterinary specialists due to the emergence of resistance of pathogenic and opportunistic fungi to antifungal agents. In recent years, information has been accumulated on the various molecular mechanisms underlying this phenomenon, but in-depth studies are needed to successfully predict resistance in various groups of fungi. To treat and prevent fungal infections several groups of antimycotics are used, where azoles and allylamines are the most frequent ones, which leads to resistance development in pathogenic and opportunistic fungi. The article presents the results of molecular methods identification of azole-resistant Candida albicans isolates and terbinafine-resistant Trichophyton isolates. The analysis of gene ERG11 nucleotide sequences of 10 Candida albicans isolates, recovered from different animal species, enabled the division of phenotypically resistant and susceptible strains, but could not differentiate between the strains, which have dose-dependent resistance to azoles. Study of single nucleotide polymorphisms in gene SQLE, associated with the resistance development to terbinafine in 12 fungal isolates of genus Trichophyton, did not allow grading them by their resistance, which is likely associated with another resistance mechanism, which can be observed in these strains. The results obtained can serve as a basis for the use of molecular methods to characterize fungi of Candida and Trichophyton genera, however, taking into account the biological features of pathogens from different groups it is reasonable to use several significant genome regions or the results of the whole genome sequencing, as well as the gene expression analysis for successful forecasting of potential resistance

Comparative study of PCR test kits for ASFV DNA detection

The paper presents comparative test results of 12 domestically produced diagnostic kits/PCR test systems for DNA detection of the African swine fever virus with regard to the following parameters: completeness and correctness of instructions for use; labeling and package contents; convenience of using the kit; shelf life stability of reagents; stability of reagents after transportation and repeated freezing – thawing; batch-to-batch repeatability; sensitivity of various test materials and specificity of kits. The study of the instructions for use and kit contents revealed incompleteness of some instructions. It was noted that some manufacturers make serious errors in the instructions, which can significantly affect the interpretation of test results. It was also observed that there is insufficient control of the manufacturing process, which results in the production of faulty kits, as well as kits with poor-quality components and errors in the labeling. Thus, during the study, one kit showed its inactivity, demonstrating the absence of accumulation curves of the fluorescent signal during amplification of both positive controls and DNA of ASFV isolates. When the specificity was assessed, all the kits showed absence of non-specific reactions and acceptable sensitivity when testing various types of ASFV-containing material (blood, suspensions of pork spleen and pork casings used in sausage production). The stability test showed a sharp deterioration in the quality of operation of one kit within the shelf life period, and a significant decrease in the fluorescence signal was detected during repeated freeze – thaw cycles for another kit. Comparison of the repeatability results of different kit batches of the same manufacturer showed significant discrepancies for 41.5% of all kits. It was found that only 33% of the studied kits for ASFV DNA detection were compliant. The results of this study demonstrate the need for control of the manufactured diagnostic kits used in state programs for animal disease monitoring

Bovine mycoplasmosis occurrence on livestock farms in the Russian Federation for 2015–2018

Mycoplasmosis control remains urgent in view of wide spread of bovine mycoplasmoses in the countries with intensive animal farming and trade relations between the Russian Federation and foreign partners including import of pedigree livestock and stud bull semen. Results of testing 1,186 biomaterial samples (blood, sera, nasal swabs, milk, preputial swabs, vaginal swabs, aborted and stillborn fetuses) collected from animals that demonstrated clinical signs of respiratory and reproductive disorders in 34 different regions of the Russian Federation for 2015–2018 are presented in the paper. The samples were tested with real-time polymerase chain reaction (rtPCR) for genomes of the following mycoplasmosis agents: Mycoplasma bovis, Mycoplasma bovigenitalium, Mycoplasma dispar. As a result, M. bovis genome was detected in 10.1% of the samples, M. bovigenitalium genome was detected in 8.6% of the samples and М. dispar genome was detected in 37.15% of the samples. Also, 927 semen samples submitted from Russian and foreign breeding farms were tested with PCR. Test results showed presence of M. bovis and M. bovigenitalium genomes in semen samples collected from native bull population. Presented data support Russian scientists’ conclusions on wide mycoplasmoses occurrence in cattle in the Russian Federation territory and risk of the disease agent introduction through semen import. All of these highlight the need for control of semen products as a source for mycoplasmosis spread as well as insufficiency of single testing of semen for granting the disease-free status to the breeding farm for genetic material marketing