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The purpose of the study was the elaboration and introduction of an effective method to confirm the identity of EAV isolates obtained in cell culture from the serum of persistently infected stallions. In view of the low cost, simplicity of preparation, and its high specificity, an indirect immunoperoxidase test based on rabbit polyclonal sera was introduced. After immunization of the rabbits with the EAV reference strain Bucyrus, purified by ultracentrifugation, a specific polyclonal serum in the titer of SN antibodies from 1:32 to 1:128 was obtained. Peroxidase-conjugated goat IgG antibodies against horse immunoglobulins were used as a secondary antibody and DAB as a substrate. After the optimalization of the test conditions, its sensitivity and specificity was defined. The sensitivity estimated in reference to the Bucyrus strain was equal to 1 TCID₅₀. The specificity was defined at 100%. After the preliminary validation, the indirect immunoperoxidase test based on the obtained polyclonal sera was affirmed as a method to identify EAV isolates. All 68 EAV field isolates reacted positively in the immunoperoxidase test.
The paper presents the current state of knowledge on the prevalence and risk of a new type of bovine viral diarrhea virus known as type 3 (BVDV-3). The first discovered atypical pestivirus was the isolate D32/00_’HOBI’, detected in fetal calf serum (FCS) originating from Brazil. The isolates CH-KaHO/cont, SVA/cont-08 and IZSPLV_To are further examples confirming the presence of BVDV type 3 in the FCS. This new species of pestiviruses (BVDV-3) is a problem not only for research laboratories using bovine serum, but also for cattle breeders. Natural infections with this virus have been reported in Brazil, Thailand and Italy, which may suggest that the new species of pestivirus is also present in European cattle. The methods used for routine diagnosis of BVDV infection are ineffective in detecting atypical pestiviruses, which may pose a risk of false negative results. It may also influence the safety of vaccines and biological products produced on the basis of contaminated batches of fetal bovine serum.
Spring viremia of carp (SVC) is a disease caused by a virus belonging to the genus Vesiculovirus, family Rhabdoviridae. The SVC virus is divided into four genogroups, Ia, Ib, Ic, and Id, due to its geographical distribution. This study aimed to identify the genotype of the SVC virus circulating in Poland. Polish SVC virus isolates were propagated on EPC and FHM cell lines, and genetic material (RNA) was isolated. The virus was detected in test samples by reverse transcription, sequenced and analyzed using MEGA 6.06 software. The phylogenetic tree was constructed by the Neighbor-Joining method. The results of phylogenetic analysis revealed the presence of two genogroups of the SVC virus in Poland. Most of Polish isolates belonged to the genogroup Id, as do isolates AY196200 from the Czech Republic, Z37505 from Belgium and EF593149 from the United States. Only two Polish isolates from Silesian Voivodeship were more closely related to Chinese and US isolates belonging to the genogroup Ia. There were no isolates belonging to the genogroups Ib and Ic. Nucleotide sequence analysis revealed certain point mutations between particular isolates. Knowledge on the genetic variants of the SVC virus circulating in Poland will be useful in epizootic investigations and preventive measures to protect Polish aquaculture from new variants from the neighboring countries.
Tomato torrado virus is a member of Secoviridae family, genus Torradovirus. It is a dangerous pathogen of tomato plants causing intensive necrosis of leaves and fruits, leading to plant death, and significantly decreasing production of this vegetable. In Poland three isolates of ToTV were identified: Wal’03, Kra and Ros. On the basis of the previously performed molecular and genomic analyses distinctive genetic variability was revealed within 3`UTR region of RNA1 Kra and Wal’03 isolates. On the basis of this heterogenous region a rapid protocol of PCR-HRM reaction was developed allowing the identification and differentiation of two isolates of Tomato torrado virus as well as constituting rapid test to monitor the nucleotide point mutation within this regulatory region. Since the 3`UTR region is known to play a role in the replication process hence the featured heterogeneity might have an impact on the control of viral particles accumulation.
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