Budachetri, K. | Search bioRxiv

bioRxiv2024

Analysis of Amblyomma americanum microRNAs in response to Ehrlichia chaffeensis infection and their potential role in vectorial capacity

BackgroundMicroRNAs (miRNAs) represent a subset of small noncoding RNAs and carry tremendous potential for regulating gene expression at the post-transcriptional level. They play pivotal roles in distinct cellular mechanisms including inhibition of bacterial, parasitic, and viral infections via immune response pathways. Intriguingly, pathogens have developed strategies to manipulate the hosts miRNA profile, fostering environments conducive to successful infection. Therefore, changes in an arthropod hosts miRNA profile in response to pathogen invasion could be critical in understanding host-pathogen dynamics. Additionally, this area of study could provide insights into discovering new targets for disease control and prevention. The main objective of the present study is to investigate the functional role of differentially expressed miRNAs upon Ehrlichia chaffeensis, a tick-borne pathogen, infection in tick vector, Amblyomma americanum. MethodsSmall RNA libraries from uninfected and E. chaffeensis-infected Am. americanum midgut and salivary gland tissues were prepared using the Illumina Truseq kit. Small RNA sequencing data was analyzed using miRDeep2 and sRNAtoolbox to identify novel and known miRNAs. The differentially expressed miRNAs were validated using a quantitative PCR assay. Furthermore, a miRNA inhibitor approach was used to determine the functional role of selected miRNA candidates. ResultsThe sequencing of small RNA libraries generated >147 million raw reads in all four libraries and identified a total of >250 miRNAs across the four libraries. We identified 23 and 14 differentially expressed miRNAs in salivary glands, and midgut tissues infected with E. chaffeensis, respectively. Three differentially expressed miRNAs (miR-87, miR-750, and miR-275) were further characterized to determine their roles in pathogen infection. Inhibition of target miRNAs significantly decreased the E. chaffeensis load in tick tissues, which warrants more in-depth mechanistic studies. ConclusionsThe current study identified known and novel miRNAs and suggests that interfering with these miRNAs may impact the vectorial capacity of ticks to harbor Ehrlichia. This study identified several new miRNAs for future analysis of their functions in tick biology and tick-pathogen interaction studies.

systems biology↗

bioRxiv2021

Changes in microbial composition, diversity, and functionality in the Amblyomma maculatum microbiome following infection with Rickettsia parkeri

BackgroundTicks are the primary vectors for emerging and resurging pathogens of public health significance worldwide. Examining tick bacterial composition, diversity, and functionality across developmental stages and tissues is necessary for designing new strategies to control ticks and prevent tick-borne diseases. MethodsA high-throughput sequencing approach was used to determine the influence of blood meal and Rickettsia parkeri infection on changes in Amblyomma maculatum microbiome composition, diversity, and functionality across the developmental timeline and in different tissues. Quantitative insight into microbial ecology analysis allowed us to determine microbial population structure, composition, and diversity. A non-metric multidimensional scaling, the sparse correlations for compositional data (SparCC) module, and phylogenetic investigation of communities by reconstruction of unobserved states 2 (PICRUSt2) software were used in the assessment. ResultsThe Amblyomma maculatum microbiome comprises ten bacterial genera present across tick life cycle stages. Among the top ten bacterial genera (the core tick microbiome), Rickettsia, Francisella, and Candidatus Midichloria are the key players, with positive interactions within each developmental stage and adult tick organ tested. The bacterial abundances, based on the number of operational taxonomic units (OTUs), increase with blood meal in each stage, helping bacterial floral growth. The growth in bacterial numbers is related to highly abundant energy metabolism orthologs with blood meal, according to functional analysis. Whereas R. parkeri had a positive correlation with Candidatus Midichloria during the tick life cycle, based on the increased number of OTUs and network analysis, this was due to an increased level of metabolic activity. Interestingly, R. parkeri replaces Francisella, based on the lower level of OTUs representing Francisella in R. parkeri-infected ticks (in all stages/organs) and negatively correlated according to network and linear discriminant analysis effect size (LEfSe). ConclusionsWe found that Rickettsia and Francisella predominate in the core microbiome of the Gulf Coast tick, whereas Candidatus Midichloria and Cutibacterium levels increase with infection. Network analysis and functional annotation suggest that R. parkeri interacts positively with Candidatus Midichloria and negatively with Francisella and that metabolic profiles are upregulated with blood meal and R. parkeri infection. Overall, this is the first study to determine the combinatorial outcome of blood meal and pathogen interaction on microbiome composition over the developmental stages of Am. maculatum. This new study expands on our existing knowledge of the Am. maculatum microbiome and further highlights the need to investigate pathogen-symbiont interactions between R. parkeri and Francisella or Candidatus Midichloria to facilitate the development of strategies for controlling tick-transmitted diseases. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=146 SRC="FIGDIR/small/465777v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@1c30a2aorg.highwire.dtl.DTLVardef@1d0713borg.highwire.dtl.DTLVardef@1b9eadborg.highwire.dtl.DTLVardef@e9f5ad_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG

microbiology↗