Validation of an automatic method for reconstruction, delineation, and measurement of levator hiatus in clinical practice.

OBJECTIVES: To evaluate the concordance between an automatic software program and manual evaluation in reconstructing, delineating, and measuring the levator hiatus (LH) on maximal Valsalva maneuver. METHODS: This was a retrospective study analyzing archived raw ultrasound imaging data of 100 patients underwent transperineal ultrasound (TPUS) examination. Each data were assessed by the automatic Smart Pelvic System software program and manual evaluation. The Dice similarity index (DSI), mean absolute distance (MAD), and Hausdorff distance (HDD) were calculated to quantify delineation accuracy of LH. Agreement between automatic and manual measurement of levator hiatus area was assessed by intraclass correlation coefficient (ICC) and Bland-Altman method. RESULTS: The satisfaction rate of automatic reconstruction was 94%. Six images were recognized as unsatisfactory reconstructed images for some gas in the rectum and anal canal. Compared with satisfactory reconstructed images, DSI of unsatisfactory reconstructed images was lower, MAD and HDD were larger (p = 0.001, p = 0.001, p = 0.006, respectively). The ICC was up to 0.987 in 94 satisfactory reconstructed images. CONCLUSIONS: The Smart Pelvic System software program had good performance in reconstruction, delineation, and measurement of LH on maximal Valsalva maneuver in clinical practice, despite misidentification of the border of posterior aspect of LH due to the influence of gas in the rectum.

CircRNA-1967 participates in the differentiation of goat SHF-SCs into hair follicle lineage by sponging miR-93-3p to enhance LEF1 expression.

Circular RNAs (circRNAs), a novel class of non-coding RNAs, can interact with miRNAs through a sequence-driven sponge mechanism, thereby regulating the expression of their downstream target genes. CircRNA-1967 was found in secondary hair follicles (SHFs) of cashmere goats, but its functions are not clear. Here, we showed that both circRNA-1967 and its host gene BNC2 had significantly higher expression in SHF bulge at anagen than those at telogen of cashmere goats. Also, circRNA-1967 participates in the differentiation of SHF stem cells (SHF-SCs) into hair follicle lineage in cashmere goats. RNA pull-down assay verified that circRNA-1967 interacts with miR-93-3p. We also indicated that circRNA-1967 promoted LEF1 expression in SHF-SCs of cashmere goats. By dual-luciferase reporter analysis, we found that circRNA-1967 up-regulated LEF1 expression through the miR-93-3p-mediated pathway. The results from this study demonstrated that circRNA-1967 participated in the differentiation of goat SHF-SCs into hair follicle lineage by sponging miR-93-3p to enhance LEF1 expression. Our founding might constitute a novel pathway for revealing the potential mechanism of the differentiation of SHF-SCs into hair follicle lineage in cashmere goats. Also, these results provided a valuable basis for further enhancing the intrinsic regeneration of cashmere goat SHFs with the formation and growth of cashmere fibers.

Identification and Classification of Long Non-Coding RNAs in the Mammary Gland of the Holstein Cow.

The mammary glands, responsible for milk secretion, are regulated at a local level by various hormones, growth factors, non-coding RNAs, and other elements. Recent research has discovered the presence of lncRNAs in these glands, with suggestions that they may be essential for the maintenance and function of mammary glands. Besides directly controlling the gene and protein expression, lncRNAs are believed to play a significant part in numerous physiological and pathological processes. This study focused on examining the mammary gland tissues of Chinese Holstein cows, to identify and categorize long non-coding RNAs (lncRNAs). The research intended to distinguish lncRNAs in the mammary tissues of Holstein cows and contrast them between lactation and non-lactation periods. In this study, mammary gland tissues were sampled from three Holstein cows in early lactation (n = 3, 30 days postpartum) and non-lactation (n = 3, 315 days postpartum) on a large dairy farm in Jiangsu province. Mammary tissue samples were collected during early lactation and again during non-lactation. In total, we detected 1905 lncRNAs, with 57.3% being 500 bp and 612 intronic lncRNAs. The exon count for lncRNAs varied from 2 to 10. It was observed that 96 lncRNA expressions markedly differed between the two stages, with 83 genes being upregulated and 53 downregulated. Enrichment analysis results revealed that Gene Ontology (GO) analysis was primarily abundant in cellular processes. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated that target genes were predominantly abundant in metabolic pathways, fatty acid biosynthesis, the immune system, and glycosphingolipid biosynthesis. This study analyzed the expression profile and characteristics of lncRNAs in the mammary gland tissues of Holstein cows during both lactation and non-lactation stages, forming a foundation for further investigation into the functional roles of lncRNAs in Holstein cows throughout lactation.

Menin regulates lipid deposition in mouse hepatocytes via interacting with transcription factor FoxO1.

Non-alcoholic fatty liver disease (NAFLD) is rapidly being recognized as the leading cause of chronic liver disease worldwide. Men1, encoding protein of menin, is a key causative gene of multiple endocrine neoplasia type 1 syndrome including pancreatic tumor. It is known that insulin that secretes by endocrine tissue pancreatic islets plays a critical role in hepatic metabolism. Mouse model of hemizygous deletion of Men1 was shown to have severe hepatic metabolism disorders. However, the molecular function of menin on lipid deposition in hepatocytes needs to be further studied. Transcriptome sequencing does show that expression suppression of Men1 in mouse hepatocytes widely affect signaling pathways involved in hepatic metabolism, such as fatty acid metabolism, insulin response, glucose metabolism and inflammation. Further molecular studies indicates that menin overexpression inhibits expressions of the fat synthesis genes Srebp-1c, Fas, and Acc1, the fat differentiation genes Pparγ1 and Pparγ2, and the fat transport gene Cd36, thereby inhibiting the fat accumulation in hepatocytes. The biological process of menin regulating hepatic lipid metabolism was accomplished by interacting with the transcription factor FoxO1, which is also found to be critical for lipid metabolism. Moreover, menin responds to insulin in hepatocytes and mediates its regulatory effect on hepatic metabolism. Our findings suggest that menin is a crucial mediation factor in regulating the hepatic fat deposition, suggesting it could be a potential important therapeutic target for NAFLD.

The polymorphism of bovine Cofilin-1 gene sequence variants and association analysis with growth traits in Qinchuan cattle.

In our study, four single nucleotide polymorphisms (SNPs) were identified in exon 2 of cofilin-1 (CFL1) gene in 488 Chinese Qinchuan (QC) cattle, which included two missense mutations T 2084G and G 2107C, two synonymous mutations T 2052C and T 2169C. Further, we evaluated haplotype frequency and linkage disequilibrium (LD) coefficient of four SNPs. At SNP T 2052C, G 2107C and T 2169C, the QC cattle population belonged to intermediate genetic diversity (0.25 < PIC-value < 0.5), whereas SNP T-2084G belonged to low polymorphism (PIC-value < 0.25). Haplotype analysis showed that 6 different haplotypes (frequency > 0.03). LD analysis showed that SNP G 2107C and T 2169C, SNP G 2107C and T 2084G were high LD, respectively (r2 > 0.33). Association analysis indicated that SNP T 2052C was significantly associated with body length, chest breadth, chest depth and body mass in the QC population (p < 0.01 or p < 0.05). SNP G 2107C was significantly associated with rump length (p < 0.05). SNP T 2169C was significantly associated with chest breadth and chest depth (p < .01 or p < .05). The results of our study suggest that the CFL1 gene may be a strong candidate gene that affects growth traits in the QC cattle breeding program.

Metformin activated AMPK signaling contributes to the alleviation of LPS-induced inflammatory responses in bovine mammary epithelial cells.

BACKGROUND: Lipopolysaccharides (LPS) derived from gram-negative bacterial are often regarded as primary inducer of bovine mammary inflammation. This study evaluated the biological response of metformin activated AMPK signaling on LPS-induced inflammatory responses and metabolic changes in primary bovine mammary epithelial cells (pbMEC). The pbMEC were exposed to either 3 mmol/L Metf. for 12 h as Metf. group (Metf.) or 2 µg/mL LPS for 6 h as LPS group (LPS). Cells pretreated with 3 mmol/L metformin for 12 h followed by washing and 2 µg/mL LPS exposure for 6 h were served as ML group (ML). PBS was added to cells as the control group (Con.). RESULTS: Pre-incubation with Metf. inhibited LPS-induced expression of pro-inflammatory genes (TNF, IL1B, IL6, CXCL8, MYD88 and TLR4) and proteins (IL-1ß, TNF-α, NLRP3, Caspase1, ASC) and was accompanied by increased activation of AMPK signaling. Compared with the LPS group, phosphorylation of p65 and IκBα in the ML group were decreased and accumulation of NF-κB in the nucleus was significantly reduced by pretreatment with metformin. Metformin protects the cells from the increase of LPS-induced binding activity of NF-κB on both TNFA and IL1B promoters. Compared with the LPS group, genes (G6PC, PCK2) and proteins (SREBP1, SCD1) related to lipogenesis and carbohydrate metabolism were downregulated while catabolic ones (PPARA, ACSL1, Glut1, HK1) were upregulated in the ML group. Furthermore, increased acetylation of H3K14 by LPS challenge was reversed by pretreatment with metformin. CONCLUSION: Altogether, our results indicated that pretreatment with metformin dampens LPS-induced inflammatory responses mediated in part by AMPK/NF-κB/NLRP3 signaling and modification of histone H3K14 deacetylation and metabolic changes.

Cadmium promotes apoptosis and inflammation via the circ08409/miR-133a/TGFB2 axis in bovine mammary epithelial cells and mouse mammary gland.

Cadmium is a common environmental heavy metal pollutant that can accumulate over long periods of time and cause disease. Thus, analysis of the molecular mechanisms affected by cadmium in the body could be of great significance for the prevention and treatment of cadmium-related diseases. In this study, flow cytometry, immunofluorescence, transmission electron microscopy, H&E (Hematoxylin Eosin) staining and TUNEL (TdT-mediated dUTP Nick-End Labeling) assays were used to verify that cadmium induced apoptosis and immune responses in bovine mammary epithelial cells (BMECs) and in mouse mammary gland. Isolated BMECs cultured with or without cadmium were collected to screen miRNA (microRNA) using high-throughput sequencing. There were 42 differentially-expressed miRNAs among which 27 were upregulated and 15 downregulated including bta-miR-133a, bta-miR-23b-5p, bta-miR-29e, bta-miR-365-5p, bta-miR-615, bta-miR-7, bta-miR-11975, bta-miR-127, and bta-miR-411a. Among those, miR-133a (which can specifically target TGFB2 (Recombinant Transforming Growth Factor Beta 2) was the most significantly downregulated with a fold-change of 5.27 in BMECs cultured with cadmium. Application of the double luciferase reporter system, western blotting, and qRT-PCR (Quantitative Real-time PCR) revealed that circ08409 can directly bind to miR-133a. Experiments demonstrated that circRNA-08409 could adsorb bta-miR-133a. Both circ08409 and TGFB2 significantly increased apoptosis and altered expression level of a series of inflammatory factors in BMECs. In contrast, miR-133a decreased significantly apoptosis and inflammation in the cells. Compared with cultures receiving only cadmium, the miR-133a+cadmium cultures exhibited significant reductions in the occurrence of late apoptosis. Overall, results indicated that circ08409 could relieve the inhibitory effect of miR-133a on TGFB2 expression by combining with miR-133a and subsequently modulating cell proliferation, apoptosis and inflammation. Overall, the data suggested that the circ08409/miR-133a/TGFB2 axis might play a role in mediating the effect of cadmium on BMECs. As such, data provide novel insights into controlling hazards that cadmium could induce in the mammary gland.

AGPAT3 Gene polymorphisms are associated with milk production traits in Chinese Holstein cows.

The current study reports the identification of previously undiscovered single-nucleotide polymorphisms (SNPs) in the bovine AGPAT3 gene and further investigates their associations with milk production traits. Our results demonstrate that the major allele C of the SNP g.12264 C > T is positively correlated with test-day milk yield, protein percentage and 305-day milk yield. Importantly, in silico analysis showed that the C/T transition at this locus gives rise to two new transcription factor binding sites (TFBS), E2F1 and Nkx3-2. Polymorphism g.18658 G > A was the only SNP associated with milk urea nitrogen (MUN) with the G allele related to an increase in milk urea nitrogen as well as fat percentage. The GG genotype of SNP g.28731 A > G was associated with the highest fat and protein percentage and lowest 305-day milk yield and somatic cell score (SCS). The association between AGPAT3 locus and milk production traits could be utilized in marker-assisted selection for the genetic improvement of milk production traits and, probably in conjunction with other traits, for selection to improve fitness of dairy cattle.

Radar Signal Modulation Recognition Based on Sep-ResNet.

With the development of signal processing technology and the use of new radar systems, signal aliasing and electronic interference have occurred in space. The electromagnetic signals have become extremely complicated in their current applications in space, causing difficult problems in terms of accurately identifying radar-modulated signals in low signal-to-noise ratio (SNR) environments. To address this problem, in this paper, we propose an intelligent recognition method that combines time-frequency (T-F) analysis and a deep neural network to identify radar modulation signals. The T-F analysis of the complex Morlet wavelet transform (CMWT) method is used to extract the characteristics of signals and obtain the T-F images. Adaptive filtering and morphological processing are used in T-F image enhancement to reduce the interference of noise on signal characteristics. A deep neural network with the channel-separable ResNet (Sep-ResNet) is used to classify enhanced T-F images. The proposed method completes high-accuracy intelligent recognition of radar-modulated signals in a low-SNR environment. When the SNR is -10 dB, the probability of successful recognition (PSR) is 93.44%.

Shear wave elastography for liver fibrosis in chronic hepatitis B: Adapting the cut-offs to alanine aminotransferase levels improves accuracy.

OBJECTIVES: To determine and validate alanine aminotransferase (ALT)-adapted dual cut-offs of liver stiffness measurements (LSMs) for assessing liver fibrosis with two-dimensional shear wave elastography (2D-SWE) in patients with chronic hepatitis B (CHB) infection. METHODS: Patients with CHB infection who underwent liver biopsy to assess liver fibrosis were consecutively included. 2D-SWE confirmation thresholds with a positive likelihood ratio ≥10 and 2D-SWE exclusion thresholds with a negative likelihood ratio ≤0.1 were identified to rule in or rule out significant fibrosis and cirrhosis, respectively. RESULTS: The first 515 patients (index cohort) and the next 421 patients (validation cohort) were included in the final analysis. The low and high cut-offs to rule out and rule in patients with significant fibrosis (≥ F2) were 5.4 kPa and 9.0 kPa, respectively, in patients with ALT levels ≤ 2 × the upper limit of normal (ULN) and 7.1 kPa and 11.2 kPa in patients with ALT levels > 2 × ULN. For cirrhosis (F4), the corresponding values were 8.1 kPa and 12.3 kPa in patients with ALT levels ≤ 2 × ULN and 11.9 kPa and 24.7 kPa in patients with ALT levels > 2 × ULN. The dual cut-off values showed an overall accuracy of more than 90% for diagnosis of the presence or absence of significant fibrosis and cirrhosis in the index and validation cohorts. There were no significant differences in the accuracy values between the cohorts (all p>0.05). CONCLUSION: The ALT-adapted dual cut-offs of LSMs showed high accuracy for diagnosis of the presence or absence of significant fibrosis and cirrhosis in patients with CHB infection. KEY POINTS: • The ALT-adapted dual cut-off values of LSMs showed high accuracy for diagnosis of the presence or absence of significant fibrosis and cirrhosis. • ALT levels did not influence the overall diagnostic accuracy for predicting significant fibrosis and cirrhosis. • The ALT-adapted dual cut-offs in patients with ALT levels > 2 × ULN were markedly higher than those in patients with ALT levels ≤ 2 × ULN.

MicroRNA-145 regulates immune cytokines via targeting FSCN1 in Staphylococcus aureus-induced mastitis in dairy cows.

Dairy cow mastitis is a detrimental factor in milk quality and food safety. Mastitis generally refers to inflammation caused by infection by pathogenic microorganisms. Our studies in recent years have revealed the role of miRNA regulation in Staphylococcus aureus-induced mastitis. In the present study, we overexpressed and suppressed miR-145 to investigate the function of miR-145 in Mac-T cells. Flow cytometry, ELISA and EdU staining were used to detect changes in the secretion of several Mac-T cytokines and in cell proliferation. We found that overexpression of miR-145 in Mac-T cells significantly reduced the secretion of IL-12 and TNF-α, but increased the secretion of IFN-γ; the proliferation of bovine mammary epithelial cells was also inhibited. Using quantitative real-time PCR (qRT-PCR), Western blotting and luciferase multiplex verification techniques, we found that miR-145 targeted and regulated FSCN1. Knock-down of FSCN1 significantly increased the secretion of IL-12, while the secretion of TNF-α was significantly downregulated in Mac-T cells. Upon S. aureus infection of mammary gland tissue, the body initiated inflammatory responses; Bta-miR-145 expression was downregulated, which reduced the inhibitory effect on the FSCN1 gene; and upregulation of FSCN1 expression promoted mammary epithelial cell proliferation to allow the recovery of damaged tissue. The results of the present study will aid in understanding the immune mechanism opposing S. aureus infection in dairy cows and will provide a laboratory research basis for the prevention and treatment of mastitis.

Screening candidate microR-15a- IRAK2 regulatory pairs for predicting the response to Staphylococcus aureus-induced mastitis in dairy cows.

We established a mastitis model using exogenous infection of the mammary gland of Chinese Holstein cows with Staphylococcus aureus and extracted total RNA from S. aureus-infected and healthy mammary quarters. Differential expression of genes due to mastitis was evaluated using Affymetrix technology and results revealed a total of 1230 differentially expressed mRNAs. A subset of affected genes was verified via Q-PCR and pathway analysis. In addition, Solexa high-throughput sequencing technology was used to analyze profiles of miRNA in infected and healthy quarters. These analyses revealed a total of 52 differentially expressed miRNAs. A subset of those results was verified via Q-PCR. Bioinformatics techniques were used to predict and analyze the correlations among differentially expressed miRNA and mRNA. Results revealed a total of 329 pairs of negatively associated miRNA/mRNA, with 31 upregulated pairs of mRNA and 298 downregulated pairs of mRNA. Differential expression of miR-15a and interleukin-1 receptor-associated kinase-like 2 (IRAK2), were evaluated by western blot and luciferase reporter assays. We conclude that miR-15a and miR-15a target genes (IRAK2) constitute potential miRNA-mRNA regulatory pairs for use as biomarkers to predict a mastitis response.

2D shear wave elastography combined with MELD improved prognostic accuracy in patients with acute-on-chronic hepatitis B liver failure.

OBJECTIVES: To evaluate accuracy of two-dimensional shear wave elastography (2D SWE) and develop and validate a new prognostic score in predicting prognosis of acute-on-chronic liver failure (ACLF) patients. METHODS: From 1 October 2013 to 30 September 2015, we consecutively enrolled 290 patients, sequentially collected data (including 2D SWE, ultrasound parameters, laboratory data and prognostic scores) and recorded patients' outcome (recovering/steady or worsening) during a 90-day follow-up period. We evaluated ability of 2D SWE to predict outcomes of acute-on-chronic hepatitis B liver failure (ACLF-HBV) patients. We developed a new score (MELD-SWE, combining MELD and SWE values) for predicting mortality risk of ACLF-HBV in 179 patients in a derivation group, and validated in 111 patients. RESULTS: 2D SWE values were higher in worsening patients than recovering/steady ones (p < 0.001). Accuracy of 2D SWE in predicting outcomes of ACLF-HBV was comparable to that of the MELD score (p = 0.441). MELD-SWE showed a significantly higher prognostic value than MELD in both derivation (AUROC, 0.80 vs. 0.76, p = 0.040) and validation (AUROC, 0.87 vs. 0.82, p = 0.018) group. CONCLUSIONS: The MELD-SWE score, combining MELD and SWE values, was superior to MELD alone for outcoming prediction in patients with ACLF-HBV. KEY POINTS: • 2D SWE is a simple prognostic evaluation tool in patients with ACLF-HBV. • MELD-SWE was created in this study: 1.3×MELD + 0.3×2D SWE (kPa). • MELD-SWE score was superior to MELD alone for outcoming prediction in ACLF-HBV. • In this study, 46.8 was the optimal cut-off value of MELD-SWE score.

Genetic differentiation and phylogeny of 27 sheep populations based on structural gene loci.

To explore the genetic divergence and phylogeny of Chinese indigenous sheep breeds, in the current study, we analyzed the polymorphisms of 5 structural loci in ten sheep populations, including Sishui Fur, Sunite, Wurank, Bayinbuluke, Altay, Small-Tailed Han, Wadi, Tan, Tong and Hu sheep. The data were then compared with those from an additional 13 Asian and 4 European sheep populations acquired by the same experimental method. Based on the genetic distance and the results of a cluster analysis, we constructed the phylogenetic relationship of 27 populations. The results showed that the sheep populations in this study could be classified into four genetic groups: "Mongolian", "Tibetan", "South-Southeast Asian" and "European" sheep groups. All 10 Chinese sheep breeds belonged to the "Mongolian sheep" lineage; however, Finnish Landrace sheep and Yunnan sheep could not be classified into any of the four groups. These results could provide a good reference for the protection and utilization of primary breed resources in China and phylogenic research on Asian sheep populations.

Toll-like receptor 4 gene polymorphisms influence milk production traits in Chinese Holstein cows.

The research reported in this Research Communication aimed to describe the influence of Toll-like receptor 4 gene polymorphisms on milk production traits in Chinese Holstein cows. Toll-like receptor 4 (TLR4) is an important member of the toll-like receptor gene family that is widely found in various organisms. Since TLR4 can identify molecular patterns from various pathogenic microorganisms and induce natural and acquired immunity, it plays an important role in disease resistance in dairy cows. Two single nucleotide polymorphisms (SNPs) of TLR4 (c.-226 G > C and c.2021 C > T) that were previously found to be associated with health traits were genotyped using Sequenom MassARRAY (Sequenom Inc., San Diego, CA) for Chinese Holstein cows (n = 866). The associations between SNPs or their haplotypes and milk production traits and somatic cell count were analyzed by the generalized linear model procedure of Statistics Analysis System software (SAS). The c.-226 G > C and c.2021 C > T showed low linkage disequilibrium (r2 = 0·192). There was no association between these two SNPs and SCC, but significant effects were found for SNP c.-226 G > C on test-day milk yield, fat content, protein content, and total solid and milk urea nitrogen (P T and the SNP haplotypes on test-day milk yield, fat content, protein content, lactose content and total solids (P C was located within several potential transcription factor binding sites, including transcription factor AP-2. The polymorphisms c.-226 G > C and c.2021 C > T had significant effects on the milk production for Chinese Holstein, and these SNP could be used for molecular marker-assisted selection of milk production.

Expression profiles of miRNAs from bovine mammary glands in response to Streptococcus agalactiae-induced mastitis.

This study aimed to describe the expression profiles of microRNAs (miRNAs) from mammary gland tissues collected from dairy cows with Streptococcus agalactiae-induced mastitis and to identify differentially expressed miRNAs related to mastitis. The mammary glands of Chinese Holstein cows were challenged with Streptococcus agalactiae to induce mastitis. Small RNAs were isolated from the mammary tissues of the test and control groups and then sequenced using the Solexa sequencing technology to construct two small RNA libraries. Potential target genes of these differentially expressed miRNAs were predicted using the RNAhybrid software, and KEGG pathways associated with these genes were analysed. A total of 18 555 913 and 20 847 000 effective reads were obtained from the test and control groups, respectively. In total, 373 known and 399 novel miRNAs were detected in the test group, and 358 known and 232 novel miRNAs were uncovered in the control group. A total of 35 differentially expressed miRNAs were identified in the test group compared to the control group, including 10 up-regulated miRNAs and 25 down-regulated miRNAs. Of these miRNAs, miR-223 exhibited the highest degree of up-regulation with an approximately 3-fold increase in expression, whereas miR-26a exhibited the most decreased expression level (more than 2-fold). The RNAhybrid software predicted 18 801 genes as potential targets of these 35 miRNAs. Furthermore, several immune response and signal transduction pathways, including the RIG-I-like receptor signalling pathway, cytosolic DNA sensing pathway and Notch signal pathway, were enriched in these predicted targets. In summary, this study provided experimental evidence for the mechanism underlying the regulation of bovine mastitis by miRNAs and showed that miRNAs might be involved in signal pathways during S. agalactiae-induced mastitis.

High seroprevalence of Coxiella burnetii in dairy cattle in China.

Coxiella burnetii is the agent of Q fever, a zoonosis which occurs worldwide. As there is little reliable data on the organism in China, we investigated C. burnetii infections in dairy cattle herds around the country. Opportunistic whole blood samples were collected from 1140 dairy cattle in 19 herds, and antibodies to phase I and II C. burnetii antigens were detected using commercial ELISA kits. Seropositive cattle (381/1140, 33 %) were detected in 13 of the 15 surveyed provinces and in 16 of the 19 herds (84 %) studied. Our data indicates C. burnetii is widespread in China and that animal and human health workers should be aware of the possibility of Q fever infection in their patients.

Molecular Detection of Anaplasma spp. and Ehrlichia spp. in Ruminants from Twelve Provinces of China.

Anaplasma spp. and Ehrlichia spp. are tick-transmitted bacteria that are of significant economic importance as they can infect large and small ruminants and also people. There is little information on anaplasmosis and ehrlichiosis in ruminants in China. 16S rRNA FRET-qPCRs were used to screen convenience whole blood samples from 2,240 domestic ruminants in 12 provinces of China for Anaplasma spp. and Ehrlichia spp. Positive samples were further analyzed with a standard PCR for the gltA. Anaplasma spp. DNA was detected in the sheep (11.7%; 13/111), goats (81.8%; 219/270), cattle (13.2%; 241/1,830), and water buffaloes (6.9%; 2/29). Ehrlichia spp. DNA was detected in sheep (1.8%; 2/111), goats (1.1%; 3/270), and cattle (3.6%; 65/1830) but not in water buffaloes (0/29). Sequencing of gltA PCR products showed that A. marginale, A. ovis, Ehrlichia canis, and Ehrlichia sp. (JX629807) were present in ruminants from China, while the 16S rRNA FRET-qPCR sequence data indicated that there might also be A. platys, A. phagocytophilum, Anaplasma sp. BL126-13 (KJ410243), and Anaplasma sp. JC3-6 (KM227012). Our study shows that domestic ruminants from China are not uncommonly infected with a variety of Anaplasma spp. and Ehrlichia spp.

Association of IL8 -105G/A with mastitis somatic cell score in Chinese Holstein dairy cows.

The single nucleotide polymorphisms (SNPs) in the 5' upstream of bovine IL8 gene were investigated in 810 Chinese Holstein cows from 35 bull families in a dairy farm in Shanghai using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) technique. The Real-time PCR and Western blot were used to detect the mRNA and protein levels of genotype Chinese Holstein dairy cows. The results showed that one SNP -105G>A was detected, designating three genotypes (GG, GA and AA) with respective frequencies of 0.38, 0.46, and 0.16. The significant association of the SNP -105G>A with somatic cell score (SCS) was identified. Genotype GG had a significantly lower SCS than genotype GA or AA (P < 0.01), and the relative mRNA expression and protein level of GG was found to be the highest. These results suggest that the genotype GG may be a useful genetic marker for mastitis resistance selection and breeding in Chinese Holstein dairy cows.

Transcriptome microRNA profiling of bovine mammary glands infected with Staphylococcus aureus.

MicroRNAs are small non-coding RNA molecules that are important regulators of gene expression at the post-transcriptional level. miRNAs impact the processes of cell proliferation, differentiation and apoptosis. Thus, the regulation of miRNA expression profiles associated with mastitis will be conducive for its control. In this study, Staphylococcus aureus (S. aureus) was administered to the mammary gland of Chinese Holstein cows to construct a bacteria-type mastitis model. Total RNA was isolated from bovine mammary gland tissue samples from the S. aureus-induced mastitis group and controls. miRNAs were analyzed using Solexa sequencing and bioinformatics processing for the experimental group and control group. Two miRNA libraries were constructed respectively. A total of 370 known bovine miRNAs and 341 novel mi RNAs were detected for the S. aureus and 358 known bovine miRNAs and 232 novel miRNAs for control groups. A total of 77 miRNAs in the S. aureus group showed significant differences compared to the control group. GO (Gene Ontology) analysis showed these target genes were involved in the regulation of cells, binding, etc., while KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis showed that these genes were enriched in endocytosis, and olfactory transduction pathways involved in cancer. These results provide an experimental basis to reveal the cause and regulatory mechanism of mastitis and also suggest the potential of miRNAs to serve as biomarkers for the diagnosis of mastitis in dairy cows.