Genetic characterization of genes encoding the major surface proteins of Anaplasma marginale from cattle isolates in Thailand reveals multiple novel variants.

Bovine anaplasmosis is a serious tick-borne disease that is responsible for economic loss worldwide. The major surface proteins (MSPs), encoded by msp1 to msp5 genes of Anaplasma marginale, play an important role in host-pathogen and tick-pathogen interactions. These markers have been used for genetic characterization and phylogenetic studies. Despite domestic reports concerning suspected outbreaks of anaplasmosis in Thailand, genetic analysis of A. marginale in the country remains largely limited. Therefore, we aim to investigate the infection rate of the rickettsia organism in the Anaplasmataceae family throughout five regions of Thailand and to further characterize the key genetic markers: msp1a, msp2, and msp5 of A. marginale. From 2016 to 2021, we collected a total of 384 cattle blood samples across 18 provinces. Overall, the infection rate of the rickettsia organism in the Anaplasmataceae family was 46.1%. Over 65% of the positive samples were confirmed as A. marginale. We successfully obtained a total of 138 A. marginale msp1a (38), msp2 (79), and msp5 (21) sequences from all regions of the country. The msp1a and msp2 genes exhibit a high degree of genetic diversity, while the msp5 gene is highly conserved among the Thai isolates. Our findings regarding msp1a corroborated the genetic heterogeneity of A. marginale strains in endemic regions worldwide. Additionally, we found multiple novel variants for the first time in the current nationwide survey. We found 45 tandem repeat characters of the msp1a sequence. Among them, 24 characters were not shared with other countries. Collectively, we expanded the extent of genetic diversity in key markers; msp1a and msp2 genes, and further confirmed the previous finding that msp5 was highly conserved. The msp1a and msp2 genes could be useful for the surveillance of newly introduced strains. The current data may also be useful in designing a vaccine containing potential epitopes of different antigens in the future.

Analysis of the transcriptome of bovine endometrial cells isolated by laser micro-dissection (2): impacts of post-partum negative energy balance on stromal, glandular and luminal epithelial cells.

BACKGROUND: In post-partum dairy cows, the energy needs to satisfy high milk production induces a status of more or less pronounced Negative Energy Balance (NEB). NEB associated with fat mobilization impairs reproductive function. In a companion paper, we described constitutive gene expression in the three main endometrial cell types (stromal, glandular and luminal epithelial cells) isolated by laser capture micro-dissection (LCM) showing the specificities of their transcriptomic profiles. This study investigates the specific impact of NEB on gene expression in these cells around 80 days after parturition at day 15 of the oestrus cycle and describes their specific response to NEB. RESULTS: Following the description of their constitutive expression, the transcriptome profiles obtained by RNA sequencing of the three cells types revealed that differences related to the severity of NEB altered mainly specific patterns of expression related to individual cell types. Number of differentially expressed genes between severe NEB (SNEB) and mild NEB (MNEB) cows was higher in ST than in LE and GE, respectively. SNEB was associated with differential expression of genes coding for proteins involved in metabolic processes and embryo-maternal interactions in ST. Under-expression of genes encoding proteins with functions related to cell structure was found in GE whereas genes encoding proteins participating in pro-inflammatory pathways were over-expressed. Genes associated to adaptive immunity were under-expressed in LE. CONCLUSION: The severity of NEB after calving is associated with changes in gene expression around 80 days after parturition corresponding to the time of breeding. Specific alterations in GEs are associated with activation of pro-inflammatory mechanisms. Concomitantly, changes in the expression of genes encoding proteins involved in cell interactions and maternal recognition of pregnancy takes place in ST. The combination of these effects possibly altering the uterine environment and embryo maternal interactions may negatively influence the establishment of pregnancy.

Analysis of the transcriptome of bovine endometrial cells isolated by laser micro-dissection (1): specific signatures of stromal, glandular and luminal epithelial cells.

BACKGROUND: A number of studies have examined mRNA expression profiles of bovine endometrium at estrus and around the peri-implantation period of pregnancy. However, to date, these studies have been performed on the whole endometrium which is a complex tissue. Consequently, the knowledge of cell-specific gene expression, when analysis performed with whole endometrium, is still weak and obviously limits the relevance of the results of gene expression studies. Thus, the aim of this study was to characterize specific transcriptome of the three main cell-types of the bovine endometrium at day-15 of the estrus cycle. RESULTS: In the RNA-Seq analysis, the number of expressed genes detected over 10 transcripts per million was 6622, 7814 and 8242 for LE, GE and ST respectively. ST expressed exclusively 1236 genes while only 551 transcripts were specific to the GE and 330 specific to LE. For ST, over-represented biological processes included many regulation processes and response to stimulus, cell communication and cell adhesion, extracellular matrix organization as well as developmental process. For GE, cilium organization, cilium movement, protein localization to cilium and microtubule-based process were the only four main biological processes enriched. For LE, over-represented biological processes were enzyme linked receptor protein signaling pathway, cell-substrate adhesion and circulatory system process. CONCLUSION: The data show that each endometrial cell-type has a distinct molecular signature and provide a significantly improved overview on the biological process supported by specific cell-types. The most interesting result is that stromal cells express more genes than the two epithelial types and are associated with a greater number of pathways and ontology terms.