Lithuanian Isolates Research Articles

First molecular characterization of Sarcocystis miescheriana in wild boars (Sus scrofa) from Latvia.

Various muscle samples of wild boar (Sus scrofa) from Latvia were studied for the presence of Sarcocystis infection by means of morphological and molecular methods. Sarcocysts were detected in 122 out of 140 (87.1%) wild boar examined. According to the morphological appearance of sarcocysts, the observed cysts belonged to one morphological type and resembled Sarcocystis miescheriana. Twenty-three sarcocysts isolated from the muscles of Latvian wild boars were molecularly characterized at 18S rRNA, ITS1 and cox1. Additionally, eight sarcocysts obtained from Lithuanian wild boars were subjected to molecular analysis in order to compare intraspecific genetic variability. The amplified 18S rRNA region using newly designed primers is sufficiently variable to separate S. miecheriana from S. suihominis. All Latvian and Lithuanian isolates were confirmed belonging to S. miescheriana. No genetic variation was detected within 18S rRNA and ITS1. By contrast, the high intraspecific genetic variability of S. miescheriana was observed within cox1 since each newly obtained sequence represented a unique haplotype. The comparison made using S. miescheriana isolates from Italian and Japanese wild boar and Chinese domestic pig revealed the genetic similarity of the samples depending on their geographical distances. The current study provides the first detection of Sarcocystis infection in wild boars from Latvia and molecular characterization of S. miescheriana.

Prevalence and molecular characterization of Haemoproteus tinnunculi from falcons in Saudi Arabia

Objective:This study aimed to inspect the occurrence of Haemoproteus tinnunculi (H. tinnunculi) in falcons from the central area of Saudi Arabia.Materials and Methods:Blood samples from 100 falcons species, including 55 Falco cherrug, 22 Falco peregrinus, 13 Falco pelegrinoides, and 10 Falco rusticolus, were collected from November 2018 to April 2019 and examined for H. tinnunculi by microscopic examination and nested PCR, targeting a cytochrome b (cytb) gene.Results:The prevalence was 1% by microscopic examination. The prevalence rate of H. tinnunculi was 1% by the microscopic method and 3% by PCR. Only F. cherrug was infected. In the sequence and phylogenetic analyses, the two cytb H. tinnunculi sequences were 100% identical and closely related to the Lithuanian isolate with 99.35% identity.Conclusions:This study presents the first report of molecular detection and characterization of H. tinnunculai in F. cherrug from the Kingdom of Saudi Arabia.

Whole-genome multilocus sequence typing of closely related broiler chicken meat origin Campylobacter jejuni ST-5 isolates

Broiler chicken meat is considered as the main source of foodborne Campylobacter infection. Campylobacter jejuni multilocus sequence type ST-5 has been previously identified as predominant among Latvian and Lithuanian broiler chicken meat products and is often isolated from Lithuanian patients with Campylobacter enteric infection. The market share of Lithuanian and Latvian origin fresh broiler chicken meat sold at Estonian retail is estimated close to 50%. In this study, we used the whole-genome sequences of Latvian and Lithuanian origin C. jejuni ST-5 broiler chicken meat isolates obtained from Estonian retail. Applied genome-by-genome whole-genome MLST revealed high similarity of investigated isolates with 1589 shared alleles, of which 218 loci were polymorphic. Latvian and Lithuanian isolates had 199 to 205 intergroup differences in shared alleles, whereas intragroup isolates differed by 27 to 31 alleles. Allele differences divided the isolates into 2 distinct groups based on the country of origin, which was supported by sequence-based phylogenetic tree. The resulting geographic separation of the isolates found in this study should be carefully interpreted taking into account the small sample size.

Prevalence and molecular variability of Apple chlorotic leaf spot virus capsid protein genes in Lithuania

Apple chlorotic leaf spot virus (ACLSV) is a most common pathogen of apples in the world. The occurrence, genetic diversity, recombination patterns and mutation of ACLSV CP in apple trees 80–120 years old from old homesteads and abandoned orchards in Lithuania were investigated. Using RT-PCR, 73.8% of samples tested positive for ACLSV. Specific PCR products to virus CP gene were sequenced and analysed from 39 RNA samples of apple trees collected in different geographic locations. Heterogenic infection of ACLSV in the same apple tree was detected in 11 samples. Fifty Lithuanian sequences of ACLSV with diversity in CP gene were submitted in NCBI and compared to 28 analogous sequences by other authors. The identity of virus CP gene among Lithuanian isolates ranged 81.6–99.7% at the nucleic and 88.6–100% at the amino acids level. ACLSV isolates significantly distinguished into two clusters and seven subclusters on phylogenetic tree. Isolates related to P205, A4, B6 and GC10a were identified in Lithuania, and relationship with Mo-5 and RC was not found. Two recombinant Lithuanian isolates of ACLSV distinguished to separate subcluster R-LT. Lithuanian isolates were characterized by combination of amino acids in conservative sites. The unique single or multiple mutations in CP gene among Lithuanian isolates were observed, and substitution in conservative sites 59, 75 and 79 was also identified. The high prevalence and diversity of latent infection in old apple trees was frequently determined due to the genomic mutation and rarely due to the recombination.

Diversity of thermal ecotypes and potential pathotypes ofBacillus thuringiensissoil isolates

Ecological diversification of Bacillus thuringiensis soil isolates was examined to determine whether bacteria adapted to grow at low temperature and/or potentially pathogenic correspond to genetically distinct lineages. Altogether, nine phylogenetic lineages were found among bacilli originating from North-Eastern Poland (n = 24) and Lithuania (n = 25) using multi-locus sequence typing. This clustering was chiefly confirmed by pulsed-field gel electrophoresis. One third of the bacilli were found to be psychrotolerant, which strongly supports the hypothesis of the existence of thermal ecotypes among B. thuringiensis. PCR screening was also performed to detect potential enterotoxin genes and Bacillus anthracis pXO1- and pXO2-like replicons. The cytK-positive isolates (22%) were significantly associated with two phylogenetic lineages (potential CytK pathotypes), whereas there was no correlation between phylogenetic grouping and the presence of the potential tripartite enterotoxin pathotypes (86% of strains). A statistically significant association between phylogenetic lineages and ecologic properties was found with regard to the cry1-positive Lithuanian isolates, while the cry genes in Polish isolates and the pXO1- and pXO2 replicon-like elements showed scattered distribution across phylogenetic lineages. Our results support the hypothesis that B. thuringiensis comprises strains belonging to different phylogenetic lineages, which exhibit specific ecological properties.

Tick-borne encephalitis virus in Lithuania.

Tick-borne encephalitis (TBE) is a severe problem in Lithuania, indicated by the 171 to 645 serologically confirmed cases that occurred each year between 1993 and 1999. In the present report, the first isolation and partial genetic analysis of a Lithuanian TBE virus (TBEV) strain isolated from a patient's serum sample is described. The patient was bitten by a tick while visiting the Lazdijai district (Veisiejai forest) in the southernmost part of Lithuania, a geographical area where Ixodes ricinus but not Ixodes persulcatus ticks are known to be present. The E protein-encoding viral gene sequence (nt 74-1273) recovered from the TBEV isolate showed the closest similarity to previously characterized European strains of the Western TBEV subtype, including the prototype TBEV strain Neudoerfl and those from neighbouring Latvia. Accordingly, the Lithuanian isolate was placed within the Western genetic lineage of TBEV in phylogenetic trees.