Pathogens like bacteria and protozoa, which affect human and animal health worldwide, can be transmitted by vectors like ticks. To investigate the epidemiology and genetic diversity of bacteria and protozoans carried by ticks in Chengmai county of Hainan province, China, 285 adult hard ticks belonging to two species [
Anaplasmataceae/isolation & purification* ; Animals ; Chaperonin 60/genetics* ; China ; Citrate (si)-Synthase/genetics* ; Coccidia/isolation & purification* ; Coxiellaceae/isolation & purification* ; Insect Vectors/microbiology* ; Islands ; Ixodidae/microbiology* ; Phylogeny ; Piroplasmia/isolation & purification* ; RNA, Ribosomal, 16S/genetics* ; RNA, Ribosomal, 18S/genetics*

Avian trematodes, Urogonimus turdi (Digenea: Leucochloridiidae), were collected from the intestine of wild birds, Zoothera aurea, 2013–2017 in the Daejeon Metropolitan City, Korea. The body was ellipsoidal, attenuated and/or round ends, 1,987–2,120 long and 819–831 μm wide. The oral sucker was subterminal, rounded anteriorly, and 308–425×351–432 μm in size; the prepharynx and esophagus were almost lacking; pharynx was well-developed, 142–179×78–170 μm in size; intestine narrow, bifurcating just after pharynx, ascending to the oral sucker before looping posteriorly and terminating near the posterior end; ventral sucker larger, in almost median, 536–673×447–605 μm and approximately 1.5 times larger than oral sucker. A phylogenetic tree constructed with 18S ribosomal RNA showed inter- and intraspecific relationships. Based on these morphological and molecular findings, we report here a U. turdi from White’s thrushes in Korea.
Birds ; Candidiasis, Oral ; Esophagus ; Intestines ; Korea ; Pharynx ; Republic of Korea ; RNA, Ribosomal, 18S ; Songbirds ; Trees

Objective To detect the diatom population diversity in Dianchi by constructing a 18S rDNA clone library. Methods DNA from diatoms in 6 water samples of Dianchi was amplified with diatom 18S rDNA specific primer.The 18S rDNA clone library was constructed, and clones were randomly selected for sequence. Sequence alignment was performed by BLAST. The diatom population distribution in Dianchi was analyzed and the phylogenetic tree of diatom 18S rDNA in Dianchi waters was established with the MEGA v7.0.14 software. Results Two hundred and forty clones were sequenced, with 167 diatom sequences obtained, including 11 diatom species such as Stephanodiscus, Diatoma, and Melosira. There were certain differences in diatom population distribution among the 6 samples. Conclusion The population distribution of diatom species in Dianchi shows unique features and the sequence analysis of diatom 18S rDNA has a certain reference value to the inference of forensic drowning sites.
China ; DNA, Ribosomal/genetics* ; Diatoms/classification* ; Drowning ; Forensic Sciences ; Humans ; Phylogeny ; RNA, Ribosomal, 18S/genetics*

Cyclospora cayetanensis is a foodborne and waterborne pathogen that causes endemic and epidemic human diarrhea worldwide. A few epidemiological studies regarding C. cayetanensis infections in China have been conducted. During 2013, a total of 291 stool specimens were collected from patients with diarrhea at a hospital in urban Shanghai. C. cayetanensis was not detected in any of the stool specimens by traditional microscopy, whereas five stool specimens (1.72%, 5/291) were positive by PCR. These positive cases confirmed by molecular technology were all in the adult group (mean age 27.8 years; 2.94%, 5/170) with watery diarrhea. Marked infection occurred in the rainy season of May and July. Sequence and phylogenetic analyses of the partial 18S rRNA genes of C. cayetanensis isolated showed intra-species diversity of this parasite. This study showed, for the first time, that C. cayetanensis is a pathogen in outpatients with diarrhea in Shanghai, albeit at a low level. However, the transmission dynamics of this parasite in these patients remain uncertain.
Adolescent ; Adult ; China ; epidemiology ; Cyclospora ; genetics ; isolation & purification ; Cyclosporiasis ; epidemiology ; Diarrhea ; etiology ; parasitology ; Feces ; parasitology ; Female ; Humans ; Male ; Middle Aged ; Outpatients ; Phylogeny ; Polymerase Chain Reaction ; RNA, Ribosomal, 18S ; analysis ; Retrospective Studies ; Young Adult

A 3-month-old female Maltese puppy was hospitalized with persistent diarrhea in a local veterinary clinic. Blood chemistry and hematology profile were analyzed and fecal smear was examined. Diarrheal stools were examined in a diagnostic laboratory, using multiplex real-time polymerase chain reaction (PCR) against 23 diarrheal pathogens. Sequence analysis was performed using nested PCR amplicon of 18S ribosomal RNA. Coccidian oocysts were identified in the fecal smear. Although multiplex real-time PCR was positive for Cyclospora cayetanensis, the final diagnosis was Cystoisospora ohioensis infection, confirmed by phylogenetic analysis of 18S rRNA. To our knowledge, this the first case report of C. ohioensis in Korea, using microscopic examination and phylogenetic analysis.
Animals ; Chemistry ; Cyclospora ; Diagnosis ; Diarrhea ; Dogs ; Female ; Hematology ; Humans ; Infant ; Korea ; Oocysts ; Polymerase Chain Reaction ; Real-Time Polymerase Chain Reaction ; RNA, Ribosomal, 18S ; Sequence Analysis

We performed a molecular genetic study on the sequences of 18S ribosomal RNA (ITS1 region) gene in 4-day-old adult worms of Macroorchis spinulosus recovered in mice experimentally infected with metacercariae from crayfish in Jeollanam-do Province, Korea. The metacercariae were round, 180 µm in average diameter, encysted with 2 layers of thick walls, but the stylet on the oral sucker was not clearly seen. The adult flukes were oval shape, and 760-820 µm long and 320-450 µm wide, with anterolateral location of 2 large testes. The phylogenetic tree based on ITS1 sequences of 6 M. spinulosus samples showed their distinguished position from other trematode species in GenBank. The most closely resembled group was Paragonimus spp. which also take crayfish or crabs as the second intermediate host. The present study is the first molecular characterization of M. spinulosus and provided a basis for further phylogenetic studies to compare with other trematode fauna in Korea.
Animals ; DNA, Ribosomal Spacer/*genetics ; Metacercariae/classification/cytology/genetics/isolation & purification ; Mice ; Phylogeny ; RNA, Ribosomal, 18S/genetics ; Trematoda/*classification/cytology/*genetics/isolation & purification

This study aimed to develop a multiplex-touchdown PCR method to simultaneously detect 3 species of protozoan parasites, i.e., Cryptosporidium parvum, Giardia lamblia, and Cyclospora cayetanensis, the major causes of traveler’s diarrhea and are resistant to standard antimicrobial treatments. The target genes included the Cryptosporidium oocyst wall protein for C. parvum, Glutamate dehydrogenase for G. lamblia, and 18S ribosomal RNA (18S rRNA) for C. cayetanensis. The sizes of the amplified fragments were 555, 188, and 400 bps, respectively. The multiplex-touchdown PCR protocol using a primer mixture simultaneously detected protozoa in human stools, and the amplified gene was detected in >1×10³ oocysts for C. parvum, >1×10⁴ cysts for G. lamblia, and >1 copy of the 18S rRNA gene for C. cayetanensis. Taken together, our protocol convincingly demonstrated the ability to simultaneously detect C. parvum, G. lamblia, and C. cayetanenesis in stool samples.
Cryptosporidium parvum* ; Cryptosporidium* ; Cyclospora* ; Diarrhea* ; Genes, rRNA ; Giardia lamblia* ; Giardia* ; Glutamate Dehydrogenase ; Humans ; Methods ; Multiplex Polymerase Chain Reaction ; Oocysts ; Parasites ; Polymerase Chain Reaction* ; RNA, Ribosomal, 18S

OBJECTIVES: To explore the correlation between the expression levels of several RNA markers in human brain tissue and early postmortem interval （PMI）. METHODS: Twelve individuals with known PMI （range from 4.3 to 22.5 h） were selected and total RNA was extracted from brain tissue. Eight commonly used RNA markers were chosen including β-actin, GAPDH, RPS29, 18S rRNA, 5S rRNA, U6 snRNA, miRNA-9 and miRNA-125b, and the expression levels were detected in brain tissue by real-time fluorescent quantitative PCR. The internal reference markers with stable expression in early PMI were screened using geNorm software and the relationship between its expression level and some relevant factors such as age, gender and cause of death were analyzed. RNA markers normalized by internal reference were inserted into the mathematic model established by previous research for PMI estimation using R software. Model quality was judged by the error rate calculated with estimated PMI. RESULTS: 5S rRNA, miRNA-9 and miRNA-125b showed quite stable expression and their expression levels had no relation with age, gender and cause of death. The error rate of estimated PMI using β-actin was 24.6%, while GAPDH was 41.0%. CONCLUSIONS 5S rRNA, miRNA-9 and miRNA-125b are suitable as internal reference markers of human brain tissue owing to their stable expression in early PMI. The expression level of β-actin correlates well with PMI, which can be used as an additional index for early PMI estimation.
Actins/analysis* ; Autopsy ; Brain/metabolism* ; Humans ; MicroRNAs/analysis* ; Models, Theoretical ; Postmortem Changes ; RNA Stability ; RNA, Ribosomal, 18S/analysis* ; RNA, Ribosomal, 5S/analysis* ; RNA, Small Nuclear/analysis* ; Real-Time Polymerase Chain Reaction ; Software

Blastocystis sp. is a common zoonotic intestinal protozoa which has been classified into 17 subtypes (STs). A cross-sectional study was conducted to determine the prevalence and subtype distribution of Blastocystis in villagers living on the Thai-Myanmar border, where the risk of parasitic infection is high. A total of 207 stool samples were collected and DNA was extracted. PCR and sequencing using primers targeting small-subunit ribosomal RNA (SSU rRNA) gene were performed. The prevalence of Blastocystis infection was 37.2% (77/207). ST3 (19.8%; 41/207) was the predominant subtype, followed by ST1 (11.6%; 24/207), ST2 (5.3%; 11/207), and ST4 (0.5%; 1/207). A phylogenetic tree was reconstructed using the maximum likelihood (ML) method based on the Hasegawa-Kishino-Yano + G + I model. The percentage of bootstrapped trees in which the associated taxa clustered together was relatively high. Some sequences of Blastocystis positive samples (TK18, 39, 46, 71, and 90) were closely related to animals (pig and cattle) indicating zoonotic risks. Therefore, proper health education in parasitic prevention for the villagers should be promoted to improve their personal hygiene. Further longitudinal studies are required to monitor the prevalence of parasitic infections after providing health education and to investigate Blastocystis ST in animals living in these villages.
Adult ; Aged ; Animals ; Blastocystis/*classification/immunology/*isolation & purification ; Blastocystis Infections/*parasitology ; Cluster Analysis ; Cross-Sectional Studies ; DNA, Protozoan/chemistry/genetics ; DNA, Ribosomal/chemistry/genetics ; Female ; Humans ; Male ; Middle Aged ; Myanmar ; Phylogeny ; RNA, Ribosomal, 18S/genetics ; Rural Population ; Sequence Analysis, DNA ; Seroepidemiologic Studies ; *Serogroup ; Thailand ; Young Adult

Siraitia grosvenorii is a traditional Chinese medicine also as edible food. This study selected six candidate reference genes by real-time quantitative PCR, the expression stability of the candidate reference genes in the different samples was analyzed by using the software and methods of geNorm, NormFinder, BestKeeper, Delta CT method and RefFinder, reference genes for S. grosvenorii were selected for the first time. The results showed that 18SrRNA expressed most stable in all samples, was the best reference gene in the genetic analysis. The study has a guiding role for the analysis of gene expression using qRT-PCR methods, providing a suitable reference genes to ensure the results in the study on differential expressed gene in synthesis and biological pathways, also other genes of S. grosvenorii.
Cucurbitaceae ; genetics ; RNA, Ribosomal, 18S ; genetics ; Real-Time Polymerase Chain Reaction ; methods ; Reference Standards