Identification of Eimeria spp. in domestic chickens raised in alternative poultry production systems in the State of São Paulo, Brazil.

The objective of this study was to identify Eimeria spp. in alternative poultry production systems (APPS) in the State of São Paulo, Brazil. Fecal samples (168) and DNA extracted from fecal samples obtained in APPS located in different Municipalities in the State of São Paulo (93) were examined by microscopy or genera-specific PCR (ITS-1 locus). Samples positive for Eimeria spp. were examined using Eimeria lata, Eimeria nagambie, and Eimeria zaria species-specific PCR protocols (ITS-2 locus) and another E. lata-specific PCR (candidate IMP1 genomic locus) followed by molecular cloning (E. lata and E. zaria ITS-2 amplicons) and genetic sequencing. All positive DNA samples were also submitted to genera-specific nested PCR (18S rRNA gene) followed by next-generation sequencing to identify Eimeria spp. Eimeria nagambie, E. zaria, and Eimeria sp. were identified by ITS2-targeted species-specific PCRs and genetic sequencing. Next-generation sequencing identified, in order of prevalence: E. nagambie; Eimeria acervulina; Eimeria mivati; Eimeria praecox; Eimeria brunetti; Eimeria mitis; Eimeria sp.; Eimeria maxima; E. zaria, and Eimeria necatrix/tenella. Our results confirmed, for the first time in Brazil, the identification of E. nagambie, E. zaria, and Eimeria spp. ITS-2 and 18S rRNA gene sequences not yet described in Brazil.

Co-expression analysis suggests lncRNA-mRNA interactions enhance antiviral immune response during acute Chikungunya fever in whole blood of pediatric patients.

Chikungunya virus is an arbovirus that causes the neglected tropical disease chikungunya fever, common in tropical areas worldwide. There is evidence that arboviruses alter host transcriptome and modulate immune response; this modulation may involve transcriptional and post-transcriptional control mechanisms mediated by long non-coding RNAs (lncRNAs). Herein, we employed bioinformatic analysis to evaluate co-expression of lncRNAs and their putative target mRNAs in whole blood during natural Chikungunya infection in adolescent boys. Sequencing data from GSE99992 was uploaded to the Galaxy web server, where data was aligned with HISAT2, gene counts were estimated with HTSeq-count, and differential expression was run with DESeq2. After gene classification with Biomart, Pearson's correlation was applied to identify potential interactions between lncRNAs and mRNAs, which were later classified into cis and trans according to genomic location (FEELnc) and binding potential (LncTar), respectively. We identified 1,975 mRNAs and 793 lncRNAs that were differentially expressed between the acute and convalescent stages of infection in the blood. Of the co-expressed lncRNAs and mRNAs, 357 potentially interact in trans and 9 in cis; their target mRNAs enriched pathways related to immune response and viral infections. Out of 52 enriched KEGG pathways, the RIG-I like receptor signaling is enriched by the highest number of target mRNAs. This pathway starts with the recognition of viral pathogens, leading to innate immune response mediated by the production of IFN-I and inflammatory cytokines. Our findings indicate that alterations in lncRNA expression in adolescent boys, induced by acute Chikungunya infection, potentially modulate mRNAs that contribute to antiviral immune responses.

Co-expression analysis of lncRNA and mRNA suggests a role for ncRNA-mediated regulation of host-parasite interactions in primary skin lesions of patients with American tegumentary leishmaniasis.

Leishmaniasis, caused by different Leishmania species, manifests as cutaneous or visceral forms. In the American continent, the cutaneous form is called American tegumentary leishmaniasis (ATL) and is primarily caused by Leishmania (Viannia) braziliensis. Mucosal leishmaniasis (ML), the most severe form of ATL, arises in approximately 20% of patients from a primary cutaneous lesion. Evidence indicates changes in overall expression patterns of mRNAs and lncRNAs of the host in response to Leishmania infection, with the parasite capable of modulating host immune response, which may contribute to disease progression. We evaluated whether the co-expression of lncRNAs and their putative target mRNAs in primary cutaneous lesions of patients with ATL could be associated with the development of ML. Previously available public RNA-Seq data from primary skin lesions of patients infected with L. braziliensis was employed. We identified 579 mRNAs and 46 lncRNAs differentially expressed in the primary lesion that subsequently progressed to mucosal disease. Co-expression analysis revealed 1324 significantly correlated lncRNA-mRNA pairs. Among these, we highlight the positive correlation and trans-action between lncRNA SNHG29 and mRNA S100A8, both upregulated in the ML group. S100A8 and its heterodimeric partner S100A9 form a pro-inflammatory complex expressed by immune cells and seems to participate in host innate immune response processes of infection. These findings expand the knowledge of the Leishmania-host interaction and indicate that the expression of lncRNAs in the primary cutaneous lesion could regulate mRNAs and play roles in disease progression.

Co-expression analysis of lncRNA and mRNA identifies potential adipogenesis regulatory non-coding RNAs involved in the transgenerational effects of tributyltin.

The obesity epidemic is considered a global public health crisis, with an increase in caloric intake, sedentary lifestyles and/or genetic predispositions as contributing factors. Although the positive energy balance is one of the most significant causes of obesity, recent research has linked early exposure to Endocrine-Disrupting Chemicals (EDCs) such as the obesogen tributyltin (TBT) to the disease epidemic. In addition to their actions on the hormonal profile, EDCs can induce long-term changes in gene expression, possibly due to changes in epigenetic patterns. Long non-coding RNAs (lncRNAs) are epigenetic mediators that play important regulatory roles in several biological processes, through regulation of gene transcription and/or translation. In this study, we explored the differential expression of lncRNAs in gonadal white adipose tissue samples from adult male C57BL/6J F4 generation, female C57BL/6J offspring exposed (F0 generation) to 50 nM TBT or 0.1% DMSO (control of vehicle) via drinking water provided during pregnancy and lactation, analyzing RNA-seq data from a publicly available dataset (GSE105051). A total of 74 lncRNAs were differentially expressed (DE), 22 were up-regulated and 52 were down-regulated in the group whose F4 ancestor was exposed in utero to 50nM TBT when compared to those exposed to 0.1% DMSO (control). Regulation of DE lncRNAs and their potential partner genes in gonadal white adipose tissue of mice ancestrally exposed to EDC TBT may be related to the control of adipogenesis, as pathway enrichment analyses showed that these gene partners are mainly involved in the metabolism of lipids and glucose and in insulin-related pathways, which are essential for obesity onset and control.

Predisposing factors for increased cortisol levels in oral cancer patients.

Cancer patients may have a dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis and abnormal secretion of cortisol. Increased cortisol levels have been associated with worse prognosis in patients with different types of tumors. Although anxiety and depression can trigger an abnormal cortisol secretion, little is known regarding the influence of these emotional disorders on HPA axis dysregulation in cancer patients when evaluating together with demographic, clinicopathological and biobehavioral variables. This cross-sectional study analyzed the pre-treatment plasma cortisol levels of 133 patients with oral squamous cell carcinoma (OSCC) and its association with demographic, clinicopathological, biobehavioral and psychological variables. Plasma cortisol levels were measured by electrochemiluminescence, and anxiety and depression symptoms were assessed using Beck Anxiety Inventory (BAI) and Depression (BDI), respectively. Demographic, clinicopathological and biobehavioral data were collected from patients' medical records. Results from multivariate analysis showed that the occurrence of cancer-induced pain was predictive for higher cortisol levels (OR = 5.388, p = 0.003). Men with OSCC were 4.5 times more likely to have higher plasma cortisol levels than women (OR = 4.472, p = 0.018). The effect of sex on cortisol concentrations was lost in the adjusted model for clinical staging (OR = 2.945, p = 0.116). The absence of chronic alcohol consumption history was a protective factor for highest hormone concentrations in oral cancer patients (OR = 0.104, p = 0.004). Anxiety symptoms measured by BAI as "hands trembling" (OR = 0.192, p = 0.016) and being "nervous" (OR = 0.207, p = 0.0004) were associated with lower cortisol levels. In contrast, the feeling of "fear of losing control" was a risk factor for highest hormone concentrations (OR = 6.508, p = 0.0004). The global score and specific symptoms of depression measured by the BDI were not predictive for plasma hormone levels (p > 0.05). Together, our results show that pain, alcohol consumption and feeling fear are independent factors for increased systemic cortisol levels in patients with oral cancer. Therefore, psychological intervention, as well as control of pain and alcohol consumption, should be considered to prevent the negative effects of cortisol secretion dysregulation in cancer patients.

Effect of maternal apical periodontitis on the final step of insulin signalling and inflammatory pathway in the adult male offspring of rats.

AIM: To evaluate the final step of insulin signalling, inflammatory pathway (related to the inhibition of insulin signalling), peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α) protein content and DNA methylation in the Slc2a4 gene promoter region in the skeletal muscle of adult male offspring of rats with apical periodontitis (AP) in a single tooth or in four teeth. METHODOLOGY: Female Wistar rats were distributed into three groups: a control group, a group with one tooth with AP and a group with four teeth with AP. Thirty days after induction of AP, female rats from all groups were mated with healthy male rats. When male offspring reached 75 days of age, the following analyses were performed in the gastrocnemius muscle (GM): insulin-stimulated Akt serine and threonine phosphorylation status; NF-κB p50 and p65 subunits phosphorylation status; GLUT4, TNF-α and PGC-1α protein content by Western blotting; GLUT4 and TNF-α gene expression by real-time polymerase chain reaction (PCR); and DNA methylation in the Slc2a4 gene promoter region by restriction digestion and real-time PCR. Analysis of variance was performed, followed by Tukey's post hoc test. p values <.05 were considered to be statistically significant. RESULTS: Maternal AP in four teeth decreased insulin-stimulated Akt serine and threonine phosphorylation status, reduced GLUT4 gene expression and its protein content, and increased NF-κB p50 and p65 subunits phosphorylation status in the GM of adult offspring. There were no alterations in the parameters analysed in the GM of adult offspring of rats with AP in a single tooth. In addition, maternal AP did not affect TNF-α gene expression and its protein content, PGC-1α protein content and DNA methylation in the Slc2a4 gene promoter region in the GM of adult offspring. CONCLUSIONS: Maternal AP in four teeth was associated with impairment in the final step of insulin signalling in the GM of adult male offspring in rats. An increase in NF-κB activity may be involved in this decrease in insulin signalling. This study demonstrates the impact of maternal AP on the health of offspring, demonstrating the importance of maintaining adequate maternal oral health to prevent diseases in adult offspring in rats.

Single nucleotide polymorphisms affect miRNA target prediction in bovine.

Single nucleotide polymorphisms (SNPs) can have significant effects on phenotypic characteristics in cattle. MicroRNAs (miRNAs) are small, non-coding RNAs that act as post-transcriptional regulators by binding them to target mRNAs. In the present study, we scanned ~56 million SNPs against 1,064 bovine miRNA sequences and analyzed, in silico, their possible effects on target binding prediction, primary miRNA formation, association with QTL regions and the evolutionary conservation for each SNP locus. Following target prediction, we show that 71.6% of miRNA predicted targets were altered as a consequence of SNPs located within the seed region of the mature miRNAs. Next, we identified variations in the Minimum Free Energy (MFE), which represents the capacity to alter molecule stability and, consequently, miRNA maturation. A total of 48.6% of the sequences analyzed showed values within those previously reported as sufficient to alter miRNA maturation. We have also found 131 SNPs in 46 miRNAs, with altered target prediction, occurring in QTL regions. Lastly, analysis of evolutionary conservation scores for each SNP locus suggested that they have a conserved biological function through the evolutionary process. Our results suggest that SNPs in microRNAs have the potential to affect bovine phenotypes and could be of great value for genetic improvement studies, as well as production.

In silico and in vitro evaluation of primers for molecular differentiation of Leishmania species.

Leishmaniasis is a zoonotic disease caused by over 20 species of protozoan parasites of the genus Leishmania. Infection is commonly spread by sandflies and produces a wide spectrum of clinical signs and symptoms. Therefore, from an epidemiological and therapeutic standpoint, it is important to detect and differentiate Leishmania spp. The objective of this study was to combinate in silico and in vitro strategies to evaluate the analytical specificity of primers previously described in the literature. According to electronic PCR (e-PCR) analysis, 23 out of 141 pairs of primers selected through literature search matched their previously reported analytical specificity. In vitro evaluation of nine of these primer pairs by quantitative PCR (qPCR) confirmed the analytical specificity of five of them at the level of Leishmania spp., L. mexicana complex or Leishmania and Viannia subgenera. Based on these findings, the combination of e-PCR and qPCR is suggested to be a valuable approach to maximize the specificity of new primer pairs for the laboratory diagnosis of infections with Leishmania spp.

Differential expression of miRNAs in canine peripheral blood mononuclear cells (PBMC) exposed to Leishmania infantum in vitro.

Visceral Leishmaniasis (VL) is a neglected tropical disease, caused by L. infantum in the New World, where dogs are the main reservoir. These parasites can regulate host immune response through miRNA differential expression in the early stages of infection; however such early response has not yet been investigated in the canine model. PBMC from healthy dogs were exposed to L. infantum in vitro and microarray analysis showed an upregulation of miR-206, miR-302d, miR-433, miR-214, miR-493, miR-514, miR-1835, miR-210, miR-539, miR-432, miR-188, miR-345 and downregulation of miR-489 and miR-503 in comparison to non-exposed control cells, at 24 h post-exposure. In silico target prediction showed that the upregulated miRNAs target 1541 genes, which can modulate important pathways involved in the early immune responses, like the "MAPK signaling pathway", one of the most relevant pathways to Leishmania survival inside host cells. These findings shed light on parasite modulation of host immunity following Leishmania infection, which in turn can be explored for drug development.

In silico and in vitro evaluation of primers for molecular differentiation of Leishmania species / Avaliação in silico e in vitro de iniciadores para a diferenciação molecular de espécies de Leishmania

Abstract Leishmaniasis is a zoonotic disease caused by over 20 species of protozoan parasites of the genus Leishmania. Infection is commonly spread by sandflies and produces a wide spectrum of clinical signs and symptoms. Therefore, from an epidemiological and therapeutic standpoint, it is important to detect and differentiate Leishmania spp. The objective of this study was to combinate in silico and in vitro strategies to evaluate the analytical specificity of primers previously described in the literature. According to electronic PCR (e-PCR) analysis, 23 out of 141 pairs of primers selected through literature search matched their previously reported analytical specificity. In vitro evaluation of nine of these primer pairs by quantitative PCR (qPCR) confirmed the analytical specificity of five of them at the level of Leishmania spp., L. mexicana complex or Leishmania and Viannia subgenera. Based on these findings, the combination of e-PCR and qPCR is suggested to be a valuable approach to maximize the specificity of new primer pairs for the laboratory diagnosis of infections with Leishmania spp.

Resumo As leishmanioses são zoonoses causadas por mais de 20 espécies de protozoários do gênero Leishmania. As infecções são comumente disseminadas por flebotomíneos e causam um amplo espectro de manifestações clínicas. Portanto, a detecção e diferenciação de espécies de Leishmania são importantes do ponto de vista epidemiológico e terapêutico. O objetivo deste estudo foi combinar estratégias in silico e in vitro para avaliar a especificidade analítica dos primers descritos anteriormente na literatura. De acordo com a PCR eletrônica (e-PCR), 23 dos 141 pares de primers selecionados por meio de pesquisa da literatura estavam de acordo com a especificidade analítica anteriormente relatada. A avaliação in vitro de nove desses pares de primers, por PCR quantitativa (qPCR), confirmou a especificidade analítica de cinco deles ao nível de espécie de Leishmania, do complexo L. mexicana ou dos subgêneros Leishmania e Viannia. Com base nos resultados, sugere-se que a combinação de e-PCR e qPCR é uma abordagem valiosa para a validação e maximização da especificidade de novos pares de primers para o diagnóstico laboratorial de infecções com Leishmania spp.

Vitamin B12 is neuroprotective in experimental pneumococcal meningitis through modulation of hippocampal DNA methylation.

BACKGROUND: Bacterial meningitis (BM) causes apoptotic damage to the hippocampus and homocysteine (Hcy) accumulation to neurotoxic levels in the cerebrospinal fluid of children. The Hcy pathway controls bioavailability of methyl, and its homeostasis can be modulated by vitamin B12, a cofactor of the methionine synthase enzyme. Herein, the neuroprotective potential and the underlying mode of action of vitamin B12 adjuvant therapy were assessed in an infant rat model of BM. METHODS: Eleven-day old rats were intracysternally infected with Streptococcus pneumoniae serotype 3, or saline, treated with B12 or placebo, and, 24 h after infection, their hippocampi were analyzed for apoptosis in the dentate gyrus, sulfur amino acids content, global DNA methylation, transcription, and proximal promoter methylation of candidate genes. Differences between groups were compared using 2-way ANOVA followed by Bonferroni post hoc test. Correlations were tested with Spearman's test. RESULTS: B12 attenuated BM-induced hippocampal apoptosis in a Hcy-dependent manner (r = 0.80, P < 0.05). BM caused global DNA hypomethylation; however, B12 restored this parameter. Accordingly, B12 increased the methylation capacity of hippocampal cells from infected animals, as inferred from the ratio S-adenosylmethionine (SAM):S-adenosylhomocysteine (SAH) in infected animals. BM upregulated selected pro-inflammatory genes, and this effect was counteracted by B12, which also increased methylation of CpGs at the promoter of Ccl3 of infected animals. CONCLUSION: Hcy is likely to play a central role in hippocampal damage in the infant rat model of BM, and B12 shows an anti-inflammatory and neuroprotective action through methyl-dependent epigenetic mechanisms.

Gene expression is associated with virulence in murine macrophages infected with Leptospira spp.

Leptospira genus contains species that affect human health with varying degrees of pathogenicity. In this context, we aimed to evaluate the differences in the modulation of host gene expression by strains of Leptospira varying in virulence. Our data showed a high number of differentially expressed transcripts in murine macrophages following 6h of infection. Leptospira infection modulated a set of genes independently of their degree of virulence. However, pathway analysis indicated that Apoptosis, ATM Signaling, and Cell Cycle: G2/M DNA Damage Checkpoint Regulation were exclusively regulated following infection with the virulent strain. Taken together, results demonstrated that species and virulence play a role during host response to Leptospira spp in murine macrophages, which could contribute to understanding the pathogenesis of leptospirosis.

Induction of miR 21 impairs the anti-Leishmania response through inhibition of IL-12 in canine splenic leukocytes.

Visceral Leishmaniasis is a chronic zoonosis and, if left untreated, can be fatal. Infected dogs have decreased cellular immunity (Th1) and develop a potent humoral response (Th2), which is not effective for elimination of the protozoan. Immune response can be modulated by microRNAs (miRNAs), however, characterization of miRNAs and their possible regulatory role in the spleen of infected dogs have not been done. We evaluated miRNA expression in splenic leukocytes (SL) from dogs naturally infected with Leishmania infantum and developing leishmaniasis (CanL; n = 8) compared to healthy dogs (n = 4). Microarray analysis showed increased expression of miR 21, miR 148a, miR 7 and miR 615, and downregulation of miR 150, miR 125a and miR 125b. Real-time PCR validated the differential expression of miR 21, miR 148a and miR 615. Further, decrease of miR 21 in SL, by means of transfection with a miR 21 inhibitor, increased the IL-12 cytokine and the T-bet/GATA-3 ratio, and decreased parasite load on SL of dogs with CanL. Taken together, these findings suggest that L. infantum infection alters splenic expression of miRNAs and that miR 21 interferes in the cellular immune response of L. infantum-infected dogs, placing this miRNA as a possible therapeutic target in CanL.

Effect of maternal periodontitis on GLUT4 and inflammatory pathway in adult offspring.

BACKGROUND: Maternal periodontal disease leads to low birth weight (LBW), insulin resistance (IR), increased TNF-α levels, and alterations in insulin signaling in adult offspring. TNF-α has been associated with the stimulation of IKKß/NF-κB, resulting in the decreased expression of GLUT4. Another mechanism that may be involved in decreasing GLUT4 expression is DNA methylation. This study aimed to evaluate in the adult offspring of rats with periodontal disease: IR, inflammatory pathways, DNA methylation, and expression of GLUT4. METHODS: Female Wistar rats were distributed into control and experimental periodontal disease groups. Seven days after induction of periodontal disease, both groups were mated with healthy male rats. After weaning, male offspring were distributed into control offspring (CN-o) and periodontal disease offspring (PED-o) groups. Body weights were measured from 0-75 days of age. At day 75, the following were measured in the offspring: IR (HOMA-IR index); TNF-α and NF-κBp65 content in the gastrocnemius muscle (GM) by western blotting; IKKα/ß, JNK, ERK 1/2, NF-κBp65, and NF-κBp50 phosphorylation status in the GM by western blotting; DNA methylation by restriction digest and real-time PCR(qAMP); and expression of GLUT4 mRNA in the GM by real-time PCR. RESULTS: LBW, IR, increases in TNF-α, IKKα/ß, ERK 1/2, NF-κBp65, and NF-κBp50 decreased expression of GLUT4 mRNA were observed in the PED-o rats. No differences were identified in JNK phosphorylation status and DNA methylation in the evaluated regions of the GLUT4-encoding gene Slc2a4. CONCLUSION: Maternal periodontal disease causes LBW, IR, activation of inflammatory pathways, and decreased GLUT4 expression in the GM of adult offspring.

Association of Copy Number Variation at Intron 3 of HMGA2 With Navel Length in Bos indicus.

Navel injuries caused by friction against the pasture can promote infection, reproductive problems and costly treatments in beef cattle raised in extensive systems. A haplotype-based genome-wide association study (GWAS) was performed for visual scores of navel length at yearling in Nellore cattle (Bos indicus) using data from 2,016 animals and 503,088 single nucleotide polymorphism (SNP) markers. The strongest signal (p = 1.01 × 10-9) was found on chromosome 5 spanning positions 47.9-48.2 Mbp. This region contains introns 3 and 4 and exons 4 and 5 of the high mobility group AT-hook 2 gene (HMGA2). Further inspection of the region with whole genome sequence data of 21 Nellore bulls revealed correlations between counts of the significant haplotype and copy number gains of a ∼6.2 kbp segment of intron 3 of HMGA2. Analysis of genome sequences from five African B. indicus and four European Bos taurus breeds revealed that the copy number variant (CNV) is indicine-specific. This intronic CNV was then validated through quantitative polymerase chain reaction (qPCR) using Angus animals as copy neutral controls. Importantly, the CNV was not detectable by means of conventional SNP-based GWAS or SNP probe intensity analyses. Given that HMGA2 affects the expression of the insulin-like growth factor 2 gene (IGF2) together with the pleomorphic adenoma gene 1 (PLAG1), and that the latter has been repeatedly shown to be associated with quantitative traits of economic importance in cattle, these findings highlight the emerging role of variants impacting the insulin-like growth factor pathway to cattle breeding.

Relationship of peripheral blood mononuclear cells miRNA expression and parasitic load in canine visceral leishmaniasis.

Visceral leishmaniasis (VL) in humans is a chronic and often fatal disease if left untreated. Dogs appear to be the main reservoir host for L. infantum infection, however, in many regions other canids such as jackals, foxes, wolves and other mammals, such as hares or black rats, have been implicated as wild reservoirs. Most dogs cannot form an effective immune response against this infection, and this could be modulated by small non-coding RNAs, called microRNAs, responsible for post-transcriptional control of gene expression. Here, we evaluated the expression of miRNAs in peripheral blood mononuclear cells (PBMC) of symptomatic dogs naturally infected with Leishmania (L.) infantum (n = 10) and compared to those of healthy dogs (n = 5). Microarray analysis revealed that miR-21, miR-424, miR-194 and miR-451 had a 3-fold increase in expression, miR-192, miR-503, and miR-371 had a 2-fold increase in expression, whereas a 2-fold reduction in expression was observed for miR-150 and miR-574. Real-time PCR validated the differential expression of miR-21, miR-150, miR-451, miR-192, miR-194, and miR-371. Parasite load of PBMC was measured by real-time PCR and correlated to the differentially expressed miRNAs, showing a strong positive correlation with expression of miR-194, a regular positive correlation with miR-371 expression, and a moderate negative correlation with miR-150 expression in PBMC. These findings suggest that Leishmania infection interferes with miRNAs expression in PBMC, and their correlation with parasite load may help in the identification of therapeutic targets in Canine Visceral Leishmaniasis (CVL).

Characterization of the microtranscriptome of macrophages infected with virulent, attenuated and saprophyte strains of Leptospira spp.

Leptospirosis is a bacterial zoonosis, caused by Leptospira spp., that leads to significant morbidity and mortality worldwide. Despite considerable advances, much is yet to be discovered about disease pathogenicity. The influence of epigenetic mechanisms, particularly RNA-mediated post-transcriptional regulation of host immune response has been described following a variety of bacterial infections. The current study examined the microtranscriptome of macrophages J774A.1 following an 8h infection with virulent, attenuated and saprophyte strains of Leptospira. Microarray analysis revealed that 29 miRNAs were misregulated following leptospiral infection compared to control macrophages in a strain and virulence-specific manner. Pathway analysis for targets of these differentially expressed miRNAs suggests that several processes involved in immune response could be regulated by miRNAs. Our data provides the first evidence that host miRNAs are regulated by Leptospira infection in macrophages. A number of the identified miRNA targets participate in key immune response processes. We suggest that post-transcriptional regulation by miRNAs may play a role in host response to infection in leptospirosis.

Data on differentially expressed miRNAs in dogs infected with Leishmania infantum.

This paper contains data on differentially expressed miRNAs in peripheral blood mononuclear cells (PBMC) of dogs naturally infected by Leishmania (L.) infantum compared to healthy dogs. In recent years, studies with miRNAs have shown that these molecules play a critical role in the regulation and function of immune response.Differentially expressed miRNAs were identified by microarray, validated by real time PCR and compared with parasite load in the dogs. Targets and pathways were analyzed using the Ingenuity Pathway Analysis program.

Transcriptome datasets of macrophages infected with different strains of Leptospira spp.

The datasets reported herein provide information about microarray experiment of macrophage cell line J774A.1 infected with three different strains of Leptospira spp. Transcriptomic profiles were generated using Affymetrix® Mouse Gene 2.1 ST Array Strip. Data was normalized and statically process, p-value < 0.01, FDR < 0.05 and log2 fold change (± 2). The microarray raw data are available in Gene Expression Omnibus (GEO) under accession number GSE105141.

Changes in tri-methylation profile of lysines 4 and 27 of histone H3 in bovine blastocysts after cryopreservation.

Pregnancy rates from cryopreserved embryos remain lower than non-cryopreserved counterparts, even though these embryos appear morphologically normal. How epigenetic events, such as histone modifications, are affected by cryopreservation of embryos remains unknown. The current study evaluated the effect of conventional freezing/thawing of in vitro produced bovine blastocyst embryos on histone modifications, H3K4me3 and H3K27me3. At day 7 of in vitro culture, blastocyst stage embryos were either frozen by conventional freezing method (-0.5 °C/min in 1.5 M ethylene glycol; F/T group) or remained in culture for an additional 18 h (Ctrl). Frozen embryos were stored in liquid N2 for 14 days, thawed and placed in culture for 36 h for recovery. Control and re-expanded frozen-thawed blastocysts from both groups were fixed in 4% paraformaldehyde and stored in PBS +0.1% triton-X at 4 °C. Immunofluorescence, utilizing antibodies against H3K4me3 and H3K27me3, was conducted and staining intensity was analyzed as percentage of total DNA. Day 7 blastocyst development rate was 35.55% (352/990) with blastocyst recovery at 54.23% (77/142) 36 h post-thawing. Total cell numbers per blastocyst were not different amongst groups (117.8 ± 12.49 and 116.1 ± 14.69, F/T and Ctrl groups respectively). Global staining for the active mark, H3K4me3, was lower in F/T blastocysts compared to Ctrl (17.24 ± 2.80% vs. 34.95 ± 3.77%; P < 0.01). However, staining for the inhibitory mark, H3K27me3, was nearly 2-fold higher in F/T blastocysts (40.41 ± 3.83% vs. 21.29 ± 3.92%; P < 0.01). These results suggest that bovine blastocysts, subjected to conventional freezing methods, have altered histone modifications that may play a role in poor pregnancy rates.