Impact factors of Blastocystis hominis infection in persons living with human immunodeficiency virus: a large-scale, multi-center observational study from China.

BACKGROUND: Blastocystis hominis (Bh) is zoonotic parasitic pathogen with a high prevalent globally, causing opportunistic infections and diarrhea disease. Human immunodeficiency virus (HIV) infection disrupts the immune system by depleting CD4+ T lymphocyte (CD4+ T) cell counts, thereby increasing Bh infection risk among persons living with HIV (PLWH). However, the precise association between Bh infection risk and HIV-related biological markers and treatment processes remains poorly understood. Hence, the purpose of the study was to explore the association between Bh infection risk and CD4+ T cell counts, HIV viral load (VL), and duration of interruption in antiviral therapy among PLWH. METHODS: A large-scale multi-center cross-sectional study was conducted in China from June 2020 to December 2022. The genetic presence of Bh in fecal samples was detected by real-time fluorescence quantitative polymerase chain reaction, the CD4+ T cell counts in venous blood was measured using flowcytometry, and the HIV VL in serum was quantified using fluorescence-based instruments. Restricted cubic spline (RCS) was applied to assess the non-linear association between Bh infection risk and CD4+ T cell counts, HIV VL, and duration of interruption in highly active antiretroviral therapy (HARRT). RESULTS: A total of 1245 PLWH were enrolled in the study, the average age of PLWH was 43 years [interquartile range (IQR): 33, 52], with 452 (36.3%) being female, 50.4% (n = 628) had no immunosuppression (CD4+ T cell counts > 500 cells/µl), and 78.1% (n = 972) achieved full virological suppression (HIV VL < 50 copies/ml). Approximately 10.5% (n = 131) of PLWH had interruption. The prevalence of Bh was found to be 4.9% [95% confidence interval (CI): 3.8-6.4%] among PLWH. Significant nonlinear associations were observed between the Bh infection risk and CD4+ T cell counts (Pfor nonlinearity < 0.001, L-shaped), HIV VL (Pfor nonlinearity < 0.001, inverted U-shaped), and duration of interruption in HARRT (Pfor nonlinearity < 0.001, inverted U-shaped). CONCLUSIONS: The study revealed that VL was a better predictor of Bh infection than CD4+ T cell counts. It is crucial to consider the simultaneous surveillance of HIV VL and CD4+ T cell counts in PLWH in the regions with high level of socioeconomic development. The integrated approach can offer more comprehensive and accurate understanding in the aspects of Bh infection and other opportunistic infections, the efficacy of therapeutic drugs, and the assessment of preventive and control strategies.

A vehicle license plate data access model based on the jump hash consistency algorithm.

The massive amount of vehicle plate data generated by intelligent transportation systems is widely used in the field of urban transportation information system construction and has a high scientific research and application value. The adoption of big data platforms to properly preserve, process, and exploit these valuable data resources has become a hot research area in recent years. To address the problems of implementing complex multi-conditional comprehensive query functions and flexible data applications in the key-value database storage environment of a big data platform, this paper proposes a data access model based on the jump hash consistency algorithm. Algorithms such as data slice storage and multi-threaded sliding window parallel reading are used to realize evenly distributed storage and fast reading of massive time-series data on clustered data nodes. A comparative analysis of data distribution uniformity and retrieval efficiency shows that the model can effectively avoid generating the cluster hotspot problem, support comprehensive analysis queries with various complex conditions, and maintain high query efficiency by precisely positioning the data storage range and utilizing parallel scan reading.

CircUBE3B High Expression Participates in Sevoflurane-Induced Human Hippocampal Neuron Injury via Targeting miR-326 and Regulating MYD88 Expression.

The clinical application of Sevoflurane (Sevo) brings about non-negligible neuron injury, leading to postoperative cognitive dysfunction (POCD). However, related pathogenesis is complex and not fully established. We aimed to disclose the role of circRNA UBE3B (circUBE3B) in neuron injury induced by Sevo. Cell viability and apoptosis were determined by CCK-8 and flow cytometry experiments. Inflammation production was monitored by ELISA. The expression of circUBE3B, miR-326, and myeloid differentiation factor 88 (MYD88) mRNA was assessed by quantitative real-time PCR (qPCR). Apoptosis-associated markers and MYD88 protein were quantified by western blot. The putative binding site between miR-326 and circUBE3B or MYD88 was verified by a dual-luciferase reporter experiment, and their binding was validated by a pull-down assay. Sevo treatment weakened cell viability and promoted cell apoptosis and inflammatory response. CircUBE3B expression was elevated in Sevo-treated neurons. Sevo-induced neuron injury was alleviated by circUBE3B downregulation but aggravated by circUBE3B overexpression. MiR-326 was targeted by circUBE3B, and miR-326 inhibition recovered neuron injury that was repressed by circUBE3B absence in Sevo-treated neurons. MiR-326 interacted with MYD88. MiR-326 enrichment attenuated Sevo-induced neuron injury, while these effects were reversed by MYD88 overexpression. CircUBE3B dysregulation was involved in Sevo-induced human hippocampal neuron injury via targeting the miR-326/MYD88 network.

Transcriptome analysis uncovers the key pathways and candidate genes related to the treatment of Echinococcus granulosus protoscoleces with the repurposed drug pyronaridine.

BACKGROUND: Cystic echinococcosis (CE) is a life-threatening zoonosis caused by the larval form of Echinococcus granulosus tapeworm. Our previous study showed that an approved drug pyronaridine (PND) is highly effective against CE, both in vitro and in an animal model. To identify possible target genes, transcriptome analysis was performed with E. granulosus sensu stricto protoscoleces treated with PND. RESULTS: A total of 1,321 genes were differentially expressed in protoscoleces treated with PND, including 541 upregulated and 780 downregulated genes. Gene ontology and KEGG analyses revealed that the spliceosome, mitogen-activated protein kinase (MAPK) pathway and ATP-binding cassette (ABC) transporters were the top three enriched pathways. Western blot analysis showed that PND treatment resulted in a dose-dependent increase in protein expression levels of EgMKK1 (MKK3/6-like) and EgMKK2 (MEK1/2-like), two members of MAPK cascades. Interestingly, several heat shock protein (HSP) genes were greatly downregulated including stress-inducible HSPs and their constitutive cognates, and some of them belong to Echinococcus-specific expansion of HSP70. CONCLUSIONS: PND has a great impact on the spliceosome, MAPK pathway and ABC transporters, which may underline the mechanisms by which PND kills E. granulosus protoscoleces. In addition, PND downregulates HSPs expression, suggesting a close relationship between the drug and HSPs.

Symptomatic and Asymptomatic Protist Infections in Hospital Inpatients in Southwestern China.

Cryptosporidium spp., Entamoeba histolytica, Giardia duodenalis, and Blastocystis sp. infections have been frequently reported as etiological agents for gastroenteritis, but also as common gut inhabitants in apparently healthy individuals. Between July 2016 and March 2017, stool samples (n = 507) were collected from randomly selected individuals (male/female ratio: 1.1, age range: 38-63 years) from two sentinel hospitals in Tengchong City Yunnan Province, China. Molecular (PCR and Sanger sequencing) methods were used to detect and genotype the investigated protist species. Carriage/infection rates were: Blastocystis sp. 9.5% (95% CI: 7.1-12.4%), G. duodenalis 2.2% (95% CI: 1.1-3.8%); and E. histolytica 2.0% (95% CI: 0.9-3.6%). Cryptosporidium spp. was not detected at all. Overall, 12.4% (95% CI: 9.7-15.6) of the participants harbored at least one enteric protist species. The most common coinfection was E. histolytica and Blastocystis sp. (1.0%; 95% CI: 0.3-2.2). Sequence analyses revealed that 90.9% (10/11) of the genotyped G. duodenalis isolates corresponded to the sub-assemblage AI. The remaining sequence (9.1%, 1/11) was identified as sub-assemblage BIV. Five different Blastocystis subtypes, including ST3 (43.7%, 21/48), ST1 (27.1%, 13/48), ST7 (18.8%, 9/48), ST4 (8.3%, 4/48), and ST2 (2.1%, 1/48) were identified. Statistical analyses confirmed that (i) the co-occurrence of protist infections was purely random, (ii) no associations were observed among the four protist species found, and (iii) neither their presence, individually or jointly, nor the patient's age was predictors for developing clinical symptoms associated with these infections. Overall, these protist mono- or coinfections are asymptomatic and do not follow any pattern.

Identification of an orally active carbazole aminoalcohol derivative with broad-spectrum anti-animal trypanosomiasis activity.

Animal trypanosomiasis, caused by the members of subgenus Trypanozoon (Trypanosoma brucei brucei, T. evansi and T. equiperdum), has reduced animal productivity leading to significant negative economic impacts in endemic regions. Due to limited drug discovery and the emergence of drug-resistance over many recent decades, novel and effective compounds against animal trypanosomiasis are urgently required. This study was conducted to evaluate the antitrypanosomal potential of a batch of carbazole aminoalcohol derivatives. Among them, we found that the most effective compound was H1402, which exhibited potent trypanocidal efficacy against the bloodstream-form of T. b. brucei (EC50 = 0.73 ± 0.05 µM) and presented low cytotoxicity against two mammalian cell lines with CC50 > 30 µM. Using a murine model of acute infection, oral administration with H1402 demonstrated a complete clearance of T. b. brucei and all the infected mice were cured when they were treated twice daily for 5 days at a dose of 100 mg/kg. Furthermore, parasites were not detected in mice infected with T. evansi and T. equiperdum (the causative agents of surra and dourine, respectively, in animals) within 30 days following the same regimen with H1402. In addition, H1402 caused severe morphological and ultrastructural destruction to trypanosomes, as well as causing phosphatidylserine externalization, which are suggested to be the most likely cause of cell death. Overall, the present data demonstrated that H1402 could be promising as a rapid, safe and orally active lead compound for the development of new chemotherapeutics for animal trypanosomiasis.

[Analysis of AVPR2 variant in a neonate with congenital nephrogenic diabetes insipidus].

OBJECTIVE: To detect potential variant in a male neonate affected with congenital nephrogenic diabetes insipidus (CNDI). METHODS: Clinical data of the patient was collected. Genomic DNA was extracted from peripheral blood samples from the child and his parents. The whole coding regions of the arginine vasopressin V2 receptor (AVPR2) gene were amplified by PCR and subjected to Sanger sequencing. RESULTS: The patient presented recurrent fever and polyuria after birth. Multiple blood gas analyses indicated hypernatremia. Ultrasound showed bilateral hydronephrosis and hydroureter. The patient was partially responsive to hydrochlorothiazide. DNA analysis identified a hemizygous frameshift variant c.890-899delACCCGGAGGC in exon 2 of the AVPR2 gene in the proband. His mother was heterozygous for the same variant. CONCLUSION: The c.890-899delACCCGGAGGC variant of the AVPR2 gene probably underlies the CNDI in the child. Above discovery has enriched to spectrum of CNDI associated variants.

Molecular Epidemiology and Risk Factors of Blastocystis sp. Infections Among General Populations in Yunnan Province, Southwestern China.

BACKGROUND: Blastocystis is a common enteric parasite of controversial pathogenic roles in human diseases. Although the prevalence of Blastocystis infections has been investigated in a diverse range of populations, there is little knowledge on the molecular epidemiology and risk factors of Blastocystis infections among general populations in southeastern China. MATERIALS AND METHODS: A total of 507 individuals were randomly selected in Yunnan province, China from July 2016 to March 2017. Stool specimens were sampled for detection of Blastocystis sp. using PCR assay, and the risk factors of Blastocystis infections were identified. Blastocystis isolates were subtyped, and the associations of Blastocystis infections and subtypes with clinical manifestations were examined. RESULTS: The overall detection rate of Blastocystis sp. was 9.47% (95% CI: 7.13-12.44%). Toilet type (OR = 3.248, 95% CI: 1.245-8.473), anemia (OR = 2.601, 95% CI: 1.245-8.473) and type of daily drinking water (OR = 3.11, 95% CI: 1.557-6.213) were identified as risk factors of Blastocystis infections; however, Blastocystis infections showed no associations with clinical symptoms. Four subtypes (ST1 to ST4) were characterized in Blastocystis isolates, in which ST3 was predominant (4.73%, 95% CI: 3.2-6.94%), followed by ST1 (3.16%, 95% CI: 1.95-5.07%), ST4 (1.38%, 95% CI: 0.07-2.82%) and ST2 (0.2%, 95% CI: 0-1.11%). In addition, ST1 subtype infection was found to correlate with anemia (OR = 4.66, 95% CI: 1.631-14.314). CONCLUSIONS: There is a high prevalence of Blastocystis infections among general populations in Yunnan province, southwestern China, and toilet type, anemia and type of daily drinking water are risk factors of Blastocystis infections. ST3 is the dominant subtype of Blastocystis sp. characterized, and ST1 correlates with anemia. Improving hygiene conditions, developing healthy lifestyles and intensifying health education programs are strongly recommended to reduce the prevalence and transmission potential of Blastocystis infections.

Methnaridine is an orally bioavailable, fast-killing and long-acting antimalarial agent that cures Plasmodium infections in mice.

BACKGROUND AND PURPOSE: Malaria is one of the deadliest diseases in the world. Novel chemotherapeutic agents are urgently required to combat the widespread Plasmodium resistance to frontline drugs. Here, we report the discovery of a novel benzonaphthyridine antimalarial, methnaridine, which was identified using a structural optimization strategy. EXPERIMENTAL APPROACH: An integrated pharmacological approach was used to evaluate the antimalarial profile of methnaridine. The pharmacokinetic properties of methnaridine were investigated along with the associated safety profile. Host immune response patterns were also analysed. KEY RESULTS: Methnaridine exhibited potent antimalarial activity against P. falciparum (3D7: IC50 = 0.0066 µM; Dd2: IC50 = 0.0056 µM). In P. berghei-infected mice, oral administration effectively suppressed parasitemia (ED50 = 0.52 mg·kg-1 ·day-1 ) and cured the established infection (CD50 = 10.13 mg·kg-1 ·day-1 ). These results are equivalent to or better than those of other antimalarial agents in clinical use. Notably, a four-dose oral regimen at a dosage of 25 mg·kg-1 achieved a complete cure of P. berghei infection in mice. Methnaridine exhibited a rapid parasiticidal profile (PCT99 = 36.0 h) and showed no cross-resistance to chloroquine. Pharmacokinetic studies revealed that methnaridine is readily absorbed, long-lasting and slowly cleared. The safety profile of methnaridine is also satisfactory (maximum tolerated dose = 1,125 mg·kg-1 ). In addition, following methnaridine treatment, infection-induced Th1 immune response was almost fully alleviated in mice. CONCLUSION AND IMPLICATIONS: Methnaridine is an orally bioavailable, fast-acting and long-lasting agent with excellent antimalarial properties. Our study highlights the potential of methnaridine for clinical development as a promising antimalarial candidate.

Study of Gilbert's Syndrome-Associated UGT1A1 Polymorphism in Jaundiced Neonates of ABO Incompatibility Hemolysis Disease.

OBJECTIVE: This study aimed to assess the probable relationship between icter in neonates with ABO incompatibility hemolysis and UGT1A1 gene polymorphism. STUDY DESIGN: There were 65 ABO hemolytic disease of the newborn (HDN) neonates of full term in the study group and 82 non-ABO HDN neonates of full term in the compared group. We tested the UGT1A1 gene mutation of neonates of ABO HDN and non-ABO HDN. We compared the incidence of hyperbilirubinemia between neonates with and without UGT1A1 mutations in the ABO HDN and non-ABO HDN, to determine the relationship between icter in neonates with ABO HDN and UGT1A1 gene polymorphism. SPSS 13.0 were used to analyze those two groups' data. RESULTS: There was statistically significant difference of the serum bilirubin level between the Gly71Arg homozygous and no mutation group in the ABO HDN patients (p < 0.05). When hyperbilirubinemia was defined as serum bilirubin concentration >342 µmol/L, the incidence of hyperbilirubinemia between patients of UGT1A1 and non-UGT1A1 mutations in the ABO HDN group was significantly different (p < 0.05). But in the non-ABO HDN group, no significant difference was found. CONCLUSION: Individuals with Gly71Arg homozygous contributed to their hyperbilirubinemia in ABO HDN patients.

[Observation on growth status of Blastocystis hominis in three media].

OBJECTIVE: To observe the growth situation of Blastocystis hominis in vitro and select the optimal method for cultivation of B. hominis in different media. METHODS: Ten positive stools with B. hominis were inoculated in three different media for cultivating, namely 1640, Jone's medium and vitro medium. And the stools with good growth status and high quantities of B. hominis were chosen to inoculate in the three media with equal amount after subcultivation, and the number of B. hominis was counted every 24 h for ten days, and the morphological changes and growth status were also observed. RESULTS: The densities of B. hominis in the 1640 and Jone's medium were higher than that in the vitro medium 48 h after the inoculation. The same stool sample was inoculated to the three different media and observed for ten days, and the results indicated that the growth of B. hominis presented regular changes in the three media, the growth peaks were on the third, sixth and ninth day post inoculation; and the density of B. hominis was the highest in the Jone's medium. The morphology of B. hominis was the clearest and most dynamic in the vitro medium, while various reproductive forms were observed in the Jone's medium. CONCLUSION: Jone's medium is suitable for the growth of B. hominis and can be the first choice for the cultivation of B. hominis in vitro, and vitro medium is the best medium for observing the growth situation of B. hominis.

A 10-year retrospective study of neonatal lupus erythematous in China.

BACKGROUND: Neonatal lupus erythematosus (NLE) is not a common disease. The death rate of complete congenital heart block (CCHB), which is the most severe clinical manifestation, is as high as 20% to 30%, so early recognition of infants at risk is important. OBJECTIVES: To investigate the clinical features and long-term prognosis of NLE. METHODS: Twenty-five cases with NLE were reviewed. The clinical manifestations of patients and their mothers were summarized and analyzed. Autoantibodies were detected, and long-term follow-up was carried out. RESULTS: There were 25 patients (male:female ratio of 11:14). CCHB was detected in only 3 of the 25 patients (12%). Cutaneous neonatal lupus erythematosus (CNLE) was seen in 22 of the 25 patients (88%). Eight babies were treated with intravenous immunoglobulin (IVIG), five of whom had a prolonged PR interval that reverted to normal sinus rhythm. During the follow-up of the patients, we found only two patients with CCHB without a pacemaker, who both exhibited growth delay. One patient with CCHB without a pacemaker died. CONCLUSIONS: Children with NLE have an excellent outcome when only skin lesions are present. Even the hepatic, hematological and neurological abnormalities are transient, with generally good outcomes. IVIG might have some effectiveness due to enhanced anti-inflammatory activity to treat early diseases that may be reversible (e.g. prolonged PR interval). The long-term prognosis for patients with NLE is still under investigation, and some infants with NLE may progress to other autoimmune diseases later in childhood.

RNA interference and the vaccine effect of a subolesin homolog from the tick Rhipicephalus haemaphysaloides.

Subolesin is a well-characterized protective antigen in many ticks and, thus, it is potentially useful in the development of a broad-spectrum vaccine or an autocidal gene silencing strategy to control tick infestations. A subolesin homolog was cloned from the tick Rhipicephalus haemaphysaloides, which is widespread in China, by rapid amplification of complementary DNA (cDNA) ends. Its full-length cDNA was 1386 base pairs (bp), containing a 483 bp open reading frame with a predicted molecular mass of 18.7 kilodaltons and an isoelectric point of 9.26. The subolesin protein had a typical nuclear localization signal in its amino-terminus. The full-length cDNA of R. haemaphysaloides showed 52 and 80% identities to those from Ixodes scapularis and R. microplus, respectively, whereas amino acid sequence alignments showed 80 and 97% identities, respectively. Native subolesin was recognized in the unfed tick midgut by an antibody against recombinant subolesin. Transcriptional analysis showed that subolesin was expressed in the tick's four developmental stages and in all of the tissues examined, except for the synganglion. The pathogen Babesia microti induced the subolesin transcript by fourfold. Subolesin gene silencing by RNA interference significantly decreased the larval engorgement rate, the attachment rate and body weight of engorged nymphs, and the body weight and attachment and engorgement rates of adults, as well as the egg weight per female tick. Vaccinating mice and rabbits with recombinant subolesin induced a significant protective effect, resulting in a reduction of blood feeding and oviposition. These results encourage further studies of using subolesin to control tick infestations in China.

[Infection of Giardia lamblia in HIV-Infected Individuals and in Kindergarden Children in Rural Area of Anhui and Genotype Analysis].

Objective: To understand the situation of Giardia lamblia infection in HIV-infected individuals and in kindergarden children in rural area of Anhui Province and analyze the genotype of the parasite. Methods: HIV-infected individuals registered in an AIDS treatment facility and children in a local kindergarden were included in this study during April 24 and May 9, 2015. The feces were collected, stained by iodine solution, and examined by microscopy. DNA was extracted from the positive feces, and nested PCR was performed to amplify the triosephosphate isomerase（tpi） gene of G. lamblia. The products were sequenced. The phylogenetic tree was constructed with BLAST, ClustalX 1.83 and MEGA6.0 softwares for analysis of homology and phylogeny. Results: One hundred and twenty-seven HIV-infected individuals and 125 kindergarden children were included. G. lamblia infection was found in three children and one HIV-infected individual. The infection detection rate in children and HIV patients was 2.40% （3/125） and 0.79% （1/127）, respectively （P>0.05）. Feces of the three infected children was soft, and no symptoms of diarrhea and stomachache were complained. Feces of the HIV-infected individual was washy, and symptoms like diarrhea, stomachache, weakness and weight loss were reported. PCR produced a specific band at 500 bp for the four persons. The sequencing results further confirmed infection in these four persons. The duplicate samples of the infected HIV patient had a 79% sequence similarity, and were 79% and 98% homologous to the Shanghai human strain of G. lamblia (GenBank accession No: KF271445), respectively. The samples of the 3 children had a 99% similarity, and all were 79% homologous to the Shanghai human strain of G. lamblia. The phylogenetic tree showed that the isolate from the HIV patient was mixed genotype of A+B, while those from the 3 children were all assemblage A. There was a high similarity between the isolates. Conclusions: There is Giardia infections in HIV patients and kindergarden children in the area. The genotype of the isolate from the HIV individual is mixed assemblage A+B while those from the children are assemblage A.

Isolation and characterization of two novel serpins from the tick Rhipicephalus haemaphysaloides.

Two novel serpins with anti-chymotrypsin activity, RHS-1 and RHS-2, were identified in the tick Rhipicephalus haemaphysaloides. The complementary cDNA sequence of RHS-1 was 1286 base pairs (bp) and encoded a deduced 403-amino acid protein with a signal peptide, whereas that of RHS-2 was 1682bp and encoded a deduced 380-amino acid protein with no signal peptide. Although both RHS-1 and RHS-2 exhibited high sequence similarities to known serpins from other ticks, the level of similarity at the amino acid level between the 2 serpins characterized here was only 32.5%. Salivary gland-specific expression of RHS-1 and midgut-specific expression of RHS-2 were found by Western blot using the relevant antiserum. We tested the ability of purified recombinant rRHS-1 and rRHS-2 to inhibit various serine proteases and found that both significantly inhibited chymotrypsin (95.6% and 94.2%, respectively). We further demonstrated that RHS-1, but not RHS-2 exhibited anticoagulation activity, based on activated partial thromboplastin time (APTT). Disruption of the genes encoding the 2 serpins with RNA interference (RNAi) led to a significant decrease in tick attachment and engorgement rates. These results indicate that RHS-1 and RHS-2 are 2 novel serpins with anti-chymotrypsin activity that are involved in blood feeding of R. haemaphysaloides.

Risk factors and clinical analysis for invasive fungal infection in neonatal intensive care unit patients.

OBJECTIVE: In this study, we seek to determine independent risk factors of invasive fungal infection (IFI) in neonatal infants. STUDY DESIGN: The medical charts of 5135 neonatal intensive care unit admissions in the past 7 years between January 2004 and December 2010 were reviewed and 45 neonates were found with IFI. Two controls, matched by gestational age, birth weight category, admission date, ward, hospital stay, and admission age, were selected for each case. RESULTS: Candida parapsilosis was the leading causative pathogen of IFI and was isolated in 33.3% of the patients. The mortality rate of the case group was 8.9% versus 1.1% in controls (p < 0.05). Multivariable logistic regression modeling defined intubation > 6 days (71.1%), use of peripherally inserted central venous catheter (68.8%), use of third-generation cephalosporin (53.3%), any prior abdominal surgeries (20.0%), and neutropenia during first week of life < 1.5 · 109/L (20.0%) as exposures significantly associated with case status. CONCLUSIONS: The predominant factors identified with IFI were third-generation cephalosporin use, peripherally inserted central venous catheter use, intubation > 6 days, any prior abdominal surgery, and neutropenia during first week of life < 1.5 · 109/L.

Distinctive microRNA profiles in the salivary glands of Haemaphysalis longicornis related to tick blood-feeding.

The salivary glands are vital to the biological success of ticks and they are a major route of pathogen transmission. Tick salivary glands undergo remarkable growth and differentiation during the blood-feeding period. MicroRNAs (miRNAs) are noncoding small RNA molecules found in diverse organisms that regulate gene expression at the post-transcriptional level. To explore transcriptional differences in the miRNAs of fed and unfed tick (Haemaphysalis longicornis) salivary glands, we investigated small RNA (sRNA) transcriptomes derived from the salivary glands and made a comparative analysis of miRNA profiles related to tick blood-feeding in the salivary glands. We generated two small RNA libraries from the salivary glands of unfed and fed H. longicornis, and obtained 14.8 and 10.3 million reads of 18-30 nt, respectively. The unfed-specific sRNAs were clearly richer than the fed-specific sRNAs in terms of the unique and total sRNAs. Overall, 769 conserved miRNA families were found in unfed samples, whereas 440 conserved miRNA families were found in fed samples. Six of the ten most abundant miRNA were found in both the unfed and fed tick salivary glands, i.e., miR-1, miR-375, bantam, miR-184, miR-739, and miR-263a. We found that known miRNA homologs displayed a wide variety of expression profiles in unfed and fed tick salivary glands. After blood-feeding, 162 known miRNAs were upregulated. The six main upregulated miRNAs were mir-1810, mir-2138, mir-2140, mir-425*, mir-429, and mir-516*. Likewise, 231 known miRNAs were downregulated after blood-feeding. The six main downregulated miRNAs were miR-2941-1*, miR-10-5p, miR-2973, miR-1183, miR-4006b-5p, and miR-881. We found that distinct microRNA profiles in the salivary glands of H. longicornis were relating to tick blood feeding. The differential expression of miRNAs in unfed and fed tick salivary glands supported their involvement at new levels in the regulation of tick blood-feeding. Our data provide an important resource for a more detailed functional analysis of miRNAs in this species.