Itemized determination of As, Sb, Bi, Se, and Ge in graphite sample by HG-AFS with high-temperature microwave digestion.

In this paper, we optimized a method for fast and accurate determination of five impurity elements (As, Sb, Bi, Se, and Ge) in graphite samples to overcome the shortcomings of existing methods, such as complicated equipment, cumbersome process, multiple-time preparation, separate determination, and large error in results. Graphite samples were digested with HNO3-H2SO4-HClO4-HF in a high-temperature and high-pressure microwave digestion apparatus, and the elements were extracted and determined separately by AFS (atomic fluorescence spectrometry). There is no element loss during the processing and analysis of this method. The spike recoveries (As: 90.30%-102.3%, Sb: 90.73%-110.0%, Bi: 90.00%-99.67%, Se: 93.33%-110.0%, Ge: 92.26%-104.2%) and precision (RSD%; As: 1.34%-8.96%, Sb: 2.67%-7.10%, Bi: 1.83%-4.58%, Se: 0.36%-3.25%, Ge: 4.41%-8.65%) meet the requirements of the corresponding quality specifications. The method has some advantages (such as no elemental loss, fast testing, strong element targeting, and accurate results), and thus can achieve batch determination of graphite samples. The optimized method for graphite sample and final solution preparations can be used for diverse spectrometric technologies, and that for spectrometer conditions have reference value for HG-AFS instruments.

Molecular characterization and zoonotic potential of Cryptosporidium spp. and Giardia duodenalis in humans and domestic animals in Heilongjiang Province, China.

BACKGROUND: Cryptosporidiosis and giardiasis are significant parasitic diseases shared between humans and domestic animals. Due to the close contact between humans and domestic animals in rural areas, it is important to consider the potential transmission of zoonotic parasites from infected domestic animals to humans. This investigation aimed to determine the prevalence and molecular characteristics of Cryptosporidium spp. and Giardia duodenalis in domestic animals and villagers. METHODS: A total of 116 fecal samples from villagers and 686 fecal samples from domestic animals in Heilongjiang Province, China, were analyzed for two parasites using nested polymerase chain reaction (PCR) targeting various genetic loci and DNA sequence analysis of the PCR products. RESULTS: By sequence analysis of the SSU rRNA gene, the prevalence of Cryptosporidium in humans was 0.9% (1/116), with one species of C. parvum (n = 1) detected; among domestic animals, the prevalence was 2.6% (18/686), with five species identified: C. suis (n = 7) and C. scrofarum (n = 7) in pigs, C. meleagridis (n = 1) in chickens, C. andersoni (n = 1) in cattle, and C. canis (n = 2) in foxes. C. parvum and C. canis were further subtyped as IIdA19G1 and XXa4 on the basis of gp60 gene. Regarding G. duodenalis, based on the SSU rRNA, bg, gdh, and tpi genes, the prevalence in domestic animals was 5.1% (31/608), with three assemblages identified: A (n = 1) in pigs, D (n = 1) in foxes, and E (n = 27) in geese, cattle, pigs, ducks, and sheep, along with mixed infection of A + E (n = 1) in one pig and B + E (n = 1) in one sheep. No G. duodenalis was detected in humans (0/116). CONCLUSIONS: The present results show that no overlap of subtypes between animals and villagers was found in Cryptosporidium spp. and G. duodenalis, indicating a minor role of domestic animals in infecting humans in this population. However, the presence of zoonotic protozoa in domestic animals highlights the need for special attention to high-risk individuals during close contact with domestic animals.

Molecular identification and genetic characteristics of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in human immunodeficiency virus/acquired immunodeficiency syndrome patients in Shanghai, China.

BACKGROUND: Opportunistic infections are a ubiquitous complication in human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) patients. Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi are common opportunistic intestinal pathogens in humans. In China, despite the number of HIV/AIDS patients being extremely large, only a few studies have investigated opportunistic infections caused by intestinal pathogens in this patient population. The aims of this study were to elucidate the occurrence and genetic characteristics of Cryptosporidium spp., G. duodenalis, and E. bieneusi in HIV/AIDS patients. METHODS: We collected fecal specimens from 155 HIV/AIDS patients (one from each patient). All of the specimens were examined for the presence of the pathogens by genotyping using polymerase chain reaction and sequencing of the small subunit ribosomal RNA gene for Cryptosporidium spp.; the triosephosphate isomerase, ß-giardin and glutamate dehydrogenase genes for G. duodenalis; and the internal transcribed spacer region of the rRNA gene for E. bieneusi. The Cryptosporidium-positive specimens were further subtyped by polymerase chain reacion and sequencing of the 60-kDa glycoprotein gene. RESULTS: Six (3.9%), three (1.9%), and eight (5.2%) HIV/AIDS patients were positive for Cryptosporidium spp., G. duodenalis, and E. bieneusi, respectively. No statistical differences were observed in occurrence rate between the groups by gender, clinical symptom (diarrhea), and CD4+ cell count. Four Cryptosporidium species were identified: Cryptosporidium hominis (n = 2), Cryptosporidium parvum (n = 1), Cryptosporidium meleagridis (n = 1), and Cryptosporidium andersoni (n = 2). Furthermore, two C. hominis subtypes (IeA12G3T3 and IaA28R4) were detected. Three G. duodenalis-positive specimens were successfully amplified and sequenced at the triosephosphate isomerase and ß-giardin loci, which led to the identification of assemblages C and B, respectively. Seven genotypes (D, Type IV, EbpC, Peru11, EbpD, A, and I) were identified in E. bieneusi-positive specimens. CONCLUSIONS: Our findings should increase awareness of AIDS-related opportunistic intestinal pathogens, and indicate the need for routine examination in clinical practice for the detection of Cryptosporidium spp., G. duodenalis, and E. bieneusi. Homology analyses of the three intestinal pathogens at the nucleotide and/or amino acid levels indicated their zoonotic potential.

Enterocytozoon bieneusi in patients with diarrhea and in animals in the northeastern Chinese city of Yichun: genotyping and assessment of potential zoonotic transmission.

Enterocytozoon bieneusi is a common microsporidia species in humans and animals. Due to lack of effective vaccines and drugs, understanding of its epidemiological status and characteristics in different hosts is an important step in controlling the infection. The present study aimed at determining the prevalence of E. bieneusi in humans with diarrhea and animals in Yichun, in northeastern China, and assessing the epidemiological role of animals in the transmission of microsporidiosis. A total of 540 fecal samples were collected from diarrheal patients (n = 222) and 11 animal species (n = 318). Enterocytozoon bieneusi was identified and genotyped by polymerase chain reaction (PCR) and sequencing of the internal transcribed spacer (ITS) region of the rRNA gene. Enterocytozoon bieneusi was detected in 1.4% (3/222) of diarrheal patients, and genotype D and novel genotypes YCHH1 and YCHH2 were identified. Enterocytozoon bieneusi was detected in wild boars (7.7%), sika deer (8.2%), dogs (3.2%), and ostriches (10.7%), and genotypes D, Type IV, Peru6, BEB6 and novel genotypes YCHA1, YCHA2 and YCHA3 were identified. Genotypes YCHH1, YCHH2 and YCHA1 were phylogenetically assigned to group 1, while YCHA2 and YCHA3 to groups 2 and 11, respectively. The finding of genotype D in humans and animals, and the identification of zoonotic genotypes Peru6, Type IV, BEB6 in animal-derived E. bieneusi isolates indicate the potential of zoonotic transmission of microsporidiosis in the investigated area. The observation of the three novel genotypes in group 1 indicates their zoonotic potential.

Title: Enterocytozoon bieneusi chez des patients souffrant de diarrhée et des animaux dans la ville de Yichun, au nord-est de la Chine: génotypage et évaluation de la transmission zoonotique potentielle. Abstract: Enterocytozoon bieneusi est une espèce de microsporidie commune chez les humains et les animaux. En raison du manque de vaccins et de médicaments efficaces, la compréhension de son statut épidémiologique et de ses caractéristiques chez différents hôtes est une étape importante dans le contrôle de l'infection. La présente étude visait à déterminer la prévalence d'E. bieneusi chez les humains souffrant de diarrhée et les animaux à Yichun, dans le nord-est de la Chine, et à évaluer le rôle épidémiologique des animaux dans la transmission de la microsporidiose. Cinq cent quarante échantillons fécaux ont été prélevés chez des patients diarrhéiques (n = 222) et 11 espèces animales (n = 318). Enterocytozoon bieneusi a été identifié et génotypé par PCR et séquençage de la région de l'espaceur interne transcrit (ITS) du gène de l'ARNr. Enterocytozoon bieneusi a été détecté chez 1,4 % (3/222) des patients souffrant de diarrhée, et le génotype D et les nouveaux génotypes YCHH1 et YCHH2 ont été identifiés. Enterocytozoon bieneusi a été détecté chez des sangliers (7,7 %), des cerfs sika (8,2 %), des chiens (3,2 %) et des autruches (10,7 %), et les génotypes D, Type IV, Peru6, BEB6 et les nouveaux génotypes YCHA1, YCHA2 et YCHA3 ont été identifiés. Les génotypes YCHH1, YCHH2 et YCHA1 ont été phylogénétiquement assignés au groupe 1, et YCHA2 et YCHA3 respectivement aux groupes 2 et 11. La découverte du génotype D chez les humains et les animaux et l'identification des génotypes zoonotiques Peru6, Type IV, BEB6 dans les isolats d'E. bieneusi d'origine animale indiquent le potentiel de transmission zoonotique de la microsporidiose dans la zone étudiée. L'observation des trois nouveaux génotypes dans le groupe 1 indique leur potentiel zoonotique.

Giardia duodenalis in patients with diarrhea and various animals in northeastern China: prevalence and multilocus genetic characterization.

BACKGROUND: Giardia duodenalis is a common parasitic diarrheal agent in humans, especially in developing countries. The aim of this study was to investigate the prevalence and multilocus genetic characterization of G. duodenalis in patients with diarrhea and animals in northeastern China, and to assess the epidemiological role of animals in the transmission of human giardiasis. METHODS: A total of 1739 fecal specimens from 413 diarrheal patients and 1326 animals comprising 16 mammal species were collected in Heilongjiang Province of China and screened for G. duodenalis by PCR and sequencing of the SSU rRNA gene. All G. duodenalis-positive specimens were subtyped by PCR and sequencing of the bg, tpi, and gdh genes. To detect additional mixed infections of different assemblages, assemblage A/B/E-specific PCRs were performed to amplify the tpi gene. RESULTS: Sequence analysis of the SSU rRNA gene determined the prevalence of G. duodenalis (5.81%, 24/413) in diarrheal patients, with a peak in minors aged 5-17 years, and identified assemblages A and B. MLG-AII and MLG-B1 were obtained based on concatenated nucleotide sequences of the bg, tpi, and gdh genes, with MLG-AII being identical to a cat-derived isolate reported previously. By sequence analysis of the SSU rRNA gene, G. duodenalis was detected in 214 (16.14%) animals belonging to 11 mammal species, with the prevalence ranging from 1.69 to 53.85%, and assemblages A to G were identified. Sequence analysis of the bg, tpi, and gdh genes from 46 specimens produced 31 MLGs, including MLG-AI (n = 1), MLG-B2-B8 (n = 18), and MLG-E1-E23 (n = 27). CONCLUSIONS: The finding of G. duodenalis in diarrheal patients enhances consciousness of detecting G. duodenalis in clinical practice and emphasizes the importance of health education in local inhabitants, especially in the age group of 5-17 years. The identification of seven assemblages (A to G) and 33 MLGs reveals genetic heterogeneity of G. duodenalis in the investigated areas. Due to insufficient homology data on the zoonotic transmission of G. duodenalis, the precise epidemiological role that animals play in the transmission of human giardiasis needs to be assessed by more large-scale molecular epidemiological investigations of local humans and animals.

Selection of reference genes for quantitative real-time PCR normalization in the coffee white stem borer, Xylotrechus quadripes Chevrolat (Coleoptera: Cerambycidae).

The coffee white stem borer, Xylotrechus quadripes Chevrolat (Coleoptera: Cerambycidae), is a major destructive pest of Coffea arabica L. (Gentianales: Rubiaceae), widely planted in many Asian countries, including China. Quantitative real-time polymerase chain reaction (qRT-PCR) is a common method for quantitative analysis of gene transcription levels. To obtain accurate and reliable qRT-PCR results, it is necessary to select suitable reference genes to different experimental conditions for normalizing the target gene expression. However, the stability of the expression of reference genes in X. quadripes has rarely been studied. In this study, the expression stability of nine candidate reference genes were investigated under biotic and abiotic conditions for use in qRT-PCR's normalization. By integrating the results of four algorithms of NormFinder, BestKeeper, geNorm, and RefFinder, the optimal reference gene combinations in different experimental conditions were performed as follows: RPL10a and EIF3D were the optimal reference genes for developmental stage samples, EIF4E, RPL10a, and RPS27a for tissue samples, V-ATP and EF1α for the sex samples, EIF3D and V-ATP for temperature treatment, RPS27a and RPL10a for insecticide stress, and RPL10a, RPS27a, and EF1α for all the samples. This study will help to obtain the stable internal reference genes under biotic and abiotic conditions and lay the foundation for in-depth functional research of target genes or genomics on olfactory molecular mechanisms, temperature adaptability, and insecticide resistance in X. quadripes.

Supramolecular structure of Artocarpus heterophyllus Lam seed starch prepared by improved extrusion cooking technology and its relationship with in vitro digestibility.

Jackfruit seed starch (JFSS) was modified by an improved extrusion cooking technology (IECT), and the supramolecular structure, molecular weight, debranched chain length distributions, relative crystallinity (Rc), and amylose content, were studied. During IECT, the α-1.4-glycosidic bond in amylopectin was broken, which led to decreased radius of gyration (Rg), number-average molar mass (Mn), weight-average molar mass (Mw), long chains and Rc. The medium and short chains and PI (Mw/Mn) increased, while the amylose content hardly changed. The crystalline structure of JFSS was converted from A-type to V-type. Increasing the temperature and screw speed during the treatment significantly increased the medium and short chains and Rg, while it decreased the long chains, amylose, Mn, Mw, PI, and Rc. However, the opposite effect was observed when increasing the moisture content. The in vitro digestibility of JFSS was significantly improved after IECT, due to destruction of starch supramolecular structure according to principal component analysis.

Lead contamination alters enzyme activities and microbial composition in the rhizosphere soil of the hyperaccumulator Pogonatherum crinitum.

Pogonatherum crinitum is a promising lead (Pb) hyperaccumulator; however, the effects of Pb contamination on P. crinitum rhizosphere soil enzymatic activities and microbial composition remain largely unexplored. Thus, an indoor experiment was conducted by cultivating P. crinitum seedlings and exposing them to four Pb concentrations (0, 1,000, 2000 and 3000 mg/kg Pb). Protease, urease, acid phosphatase and invertase activities were determined using standard methods while soil bacterial composition was determined by 16 S rDNA sequencing. The results showed that rhizosphere soil acid phosphatase activity significantly increased with increasing Pb concentration, while urease activity was significantly greater in rhizosphere soil contaminated with 1000 and 2000 mg/kg than in the control. There was a clear shift in bacterial composition during phytoremediation by P. crinitum. Compared to the control, Bacteroidetes was more abundant in all Pb-contaminated soils, Actinobacteria was more abundant in 1000 mg/kg Pb-treated soil, and Firmicutes was more abundant in 3000 mg/kg Pb-treated soil. Positive correlations were observed between dominant bacterial phyla and soil enzyme activities. Metabolic pathways, such as ABC transporter, quinine reductase, and ATP-binding protein were significantly increased in rhizosphere soil bacteria with Pb contamination. In conclusion, Pb contamination differentially influenced the activities of rhizosphere soil enzymes, specifically increasing acid phosphatase and urease activities, and alters the dominance of soil bacteria through up-regulation of genes related to some metabolic pathways. The strong correlations between dominant bacterial phyla and enzymatic activities suggest synergetic effects on the growth of P. crinitum during Pb contamination.

Prevalence and Genetic Characterization of Two Mitochondrial Gene Sequences of Strobilocercus Fasciolaris in the Livers of Brown Rats (Rattus norvegicus) in Heilongjiang Province in Northeastern China.

Rodents constitute the largest and most successful group of mammals worldwide. Brown rats (Rattus norvegicus) are one of the most common rodent species, and they serve as intermediate hosts of Hydatigera taeniaeformis. Although there have been a few studies reporting on the presence of the larval form of H. taeniaeformis (strobilocercus fasciolaris) in brown rats worldwide, little information is available on the genetic characterization of this parasite, with no molecular data from China. Therefore, from April 2014 to March 2016, this study was carried out to understand the prevalence and genetic characters of strobilocercus fasciolaris in brown rats captured in Heilongjiang Province in northeastern China. The livers of brown rats were collected and examined for the presence of cysts. Each cyst was identified based on morphological observation: the larvae with the naked eye and the scolexes under a microscope. The results were confirmed by polymerase chain reaction (PCR) and sequencing of the cytochrome c oxidase subunit 1 (cox1) and NADH dehydrogenase subunit 4 (nad4) genes. At the investigated sites, 11.8% (13/110) of the brown rats were infected with strobilocercus fasciolaris. Based on sequence analysis, there were 10 and six haplotypes regarding the cox1 and the nad4 loci, with 24 and 42 polymorphic sites, respectively (degree of intraspecific variation: 0.3%-4.4% and 0.6%-4.7%, respectively). Twelve nucleotide sequences (six of the 10 at the cox1 locus and all six at the nad4 locus) have not previously been described. Base differences in three of the six novel cox1 gene sequences and five of the six novel nad4 gene sequences caused amino acid changes. Phylogenetic analyses of the cox1 and nad4 gene sequences based on neighbor-joining and Bayesian inference trees indicated that all the strobilocercus fasciolaris isolates belonged to Hydatigera taeniaeformis sensu stricto (s.s.). This is the first report on the genetic characterization of strobilocercus fasciolaris in brown rats in China. The findings of novel cox1 and nad4 nucleotide and amino acid sequences may reflect the region-specific genetic characterization of the parasite. The data will be useful to explore the biological and epidemiological significance of the intraspecific variation within H. taeniaeformis s.s.

Identification of Uncommon Cryptosporidiumviatorum (a Novel Subtype XVcA2G1c) and Cryptosporidium andersoni as Well as Common Giardia duodenalis Assemblages A and B in Humans in Myanmar.

Cryptosporidium and Giardia are two important zoonotic intestinal protozoa responsible for diarrheal diseases in humans and animals worldwide. Feces from infected hosts, water and food contaminated by Cryptosporidium oocysts and Giardia cysts as well as predictors such as poverty have been involved in their transmission. Myanmar is one of the world's most impoverished countries. To date, there are few epidemiological studies of Cryptosporidium and Giardia in humans. To understand the prevalence and genetic characterization of Cryptosporidium spp. and Giardia duodenalis in humans in Myanmar, a molecular epidemiological investigation of the two protozoa was conducted in four villages of Shan State. 172 fecal specimens were collected from Wa people (one each) and identified for the presence of Cryptosporidium spp. and G. duodenalis by sequence analysis of their respective small subunit ribosomal RNA genes. 1.74% of investigated people were infected with Cryptosporidium spp.-C. andersoni (n = 2) and C. viatorum (n = 1) while 11.05% infected with G. duodenalis-assemblages A (n = 6) and B (n = 13). By sequence analysis of 60-kDa glycoprotein gene, the C. viatorum isolate belonged to a novel subtype XVcA2G1c. DNA preparations positive for G. duodenalis were further subtyped. Five of them were amplified and sequenced successfully: different assemblage B sequences (n = 2) at the triosephosphate isomerase (tpi) locus; sub-assemblage AII sequence (n = 1) and identical assemblage B sequences (n = 2) at the ß-giardin (bg) locus. This is the first molecular epidemiological study of Cryptosporidium spp. and G. duodenalis in humans in Myanmar at both genotype and subtype levels. Due to unclear transmission patterns and dynamics of Cryptosporidium spp. and G. duodenalis, future research effort should focus on molecular epidemiological investigations of the two parasites in humans and animals living in close contact in the investigated areas, even in whole Myanmar. These data will aid in making efficient control strategies to intervene with and prevent occurrence of both diseases.

Two lichens differing in element concentrations have similar spatial patterns of element concentrations responding to road traffic and soil input.

Two epiphytic lichens (Xanthoria alfredii, XAa; X. ulophyllodes, XAu) and soil were sampled at three sites with varied distances to a road in a semiarid sandland in Inner Mongolia, China and analyzed for concentrations of 42 elements to assess the contribution of soil input and road traffic to lichen element burdens, and to compare element concentration differences between the two lichens. The study showed that multielement patterns, Fe:Ti and rare earth element ratios were similar between the lichen and soil samples. Enrichment factors (EFs) showed that ten elements (Ca, Cd, Co, Cu, K, P, Pb, S, Sb, and Zn) were enriched in the lichens relative to the local soil. Concentrations of most elements were higher in XAu than in XAa regardless of sites, and increased with proximity to the road regardless of lichen species. These results suggested that lichen element compositions were highly affected by soil input and road traffic. The narrow-lobed sorediate species were more efficient in particulate entrapment than the broad-lobed nonsorediate species. XAa and XAu are good bioaccumulators for road pollution in desert and have similar spatial patterns of element concentrations for most elements as response to road traffic emissions and soil input.

Development of a specimen-specific in vitro pre-clinical simulation model of the human cadaveric knee with appropriate soft tissue constraints.

A human cadaveric specimen-specific knee model with appropriate soft tissue constraints was developed to appropriately simulate the biomechanical environment in the human knee, in order to pre-clinically evaluate the biomechanical and tribological performance of soft tissue interventions. Four human cadaveric knees were studied in a natural knee simulator under force control conditions in the anterior posterior (AP) and tibial rotation (TR) axes, using virtual springs to replicate the function of soft tissues. The most appropriate spring constraints for each knee were determined by comparing the kinematic outputs in terms of AP displacement and TR angle of the human knee with all the soft tissues intact, to the same knee with all the soft tissues resected and replaced with virtual spring constraints (spring rate and free length/degree). The virtual spring conditions that showed the least difference in the AP displacement and TR angle outputs compared to the intact knee were considered to be the most appropriate spring conditions for each knee. The resulting AP displacement and TR angle profiles under the appropriate virtual spring conditions all showed similar shapes to the individual intact knee for each donor. This indicated that the application of the combination of virtual AP and TR springs with appropriate free lengths/degrees was successful in simulating the natural human knee soft tissue function. Each human knee joint had different kinematics as a result of variations in anatomy and soft tissue laxity. The most appropriate AP spring rate for the four human knees varied from 20 to 55 N/mm and the TR spring rate varied from 0.3 to 1.0 Nm/°. Consequently, the most appropriate spring condition for each knee was unique and required specific combinations of spring rate and free length/degree in each of the two axes.

Multilocus Sequence Typing of Enterocytozoon bieneusi Isolates From Various Mammal and Bird Species and Assessment of Population Structure and Substructure.

Enterocytozoon bieneusi is one of the most common intestinal pathogens in humans and animals. E. bieneusi has been confirmed to be complex microsporidian species. Approximately 500 ITS genotypes of E. bieneusi have been defined. With the establishment and application of multilocus sequencing typing and population genetic tools in E. bieneusi, the studies on these aspects have been carried out worldwide, but little information is available. To understand genetic variation of mini-/micro-satellites and the population structure and substructure of E. bieneusi in northeastern China, 305 E. bieneusi DNA specimens composed of 28 ITS genotypes were from 13 mammal species and five bird species in the investigated areas. They were characterized by nested-PCR amplification and sequencing at four mini-/micro-satellite loci (MS1, MS3, MS4, and MS7). At the MS1, MS3, MS4, and MS7 loci, 153 (50.16%), 131 (42.95%), 133 (43.61%), and 128 (41.97%) DNA specimens were amplified and sequenced successfully with 44, 17, 26, and 24 genotypes being identified, respectively. Multilocus genotypes (MLGs) showed a higher genetic diversity than ITS genotypes. 48 MLGs were produced out of 90 ITS-positive DNA specimens based on concatenated sequences of all the five genetic loci including ITS. Linkage disequilibrium (LD) and limited genetic recombination were observed by measuring LD using both multilocus sequences and allelic profile data, indicating an overall clonal population structure of E. bieneusi in the investigated areas. These data will aid in the longitudinal tracking of the attribution of source of infection/contamination and in elucidating transmission dynamics, and will provide valuable information for making efficient control strategies to intervene with and prevent occurrence of microsporidiosis caused by E. bieneusi among animals and transmission of E. bieneusi from animals to humans in the investigated areas. Phylogenetic and network analyses identified three different subgroups, revealing the presence of host-shaped segregation and the absence of geographical segregation in E. bieneusi population. Meanwhile, the MLGs from zoonotic ITS genotypes were observed to be basically separated from the MLGs from host-adapted ones. Assessment of substructure will have a reference effect on understanding of zoonotic or interspecies transmission of E. bieneusi and evolution direction from zoonotic genotypes to host-adapted genotypes.

Development of robust finite element models of porcine tibiofemoral joints loaded under varied flexion angles and tibial freedoms.

The successful development of cartilage repair treatments for the knee requires understanding of the biomechanical environment within the joint. Computational finite element models play an important role in non-invasively understanding knee mechanics, but it is important to compare model findings to experimental data. The purpose of this study was to develop a methodology for generating subject-specific finite element models of porcine tibiofemoral joints that was robust and valid over multiple different constraint scenarios. Computational model predictions of two knees were compared to experimental studies on corresponding specimens loaded under several different constraint scenarios using a custom designed experimental rig, with variations made to the femoral flexion angle and level of tibial freedom. For both in vitro specimens, changing the femoral flexion angle had a marked effect on the contact distribution observed experimentally. With the tibia fixed, the majority of the contact region shifted to the medial plateau as flexion was increased. This did not occur when the tibia was free to displace and rotate in response to applied load. These trends in contact distribution across the medial and lateral plateaus were replicated in the computational models. In an additional model with the meniscus removed, contact pressures were elevated by a similar magnitude to the increase seen when the meniscus was removed experimentally. Overall, the models were able to capture specimen-specific trends in contact distribution under a variety of different loads, providing the potential to investigate subject-specific outcomes for knee interventions.

A retrospective epidemiological analysis of human Cryptosporidium infection in China during the past three decades (1987-2018).

BACKGROUND: Cryptosporidiosis is an emerging infectious disease of public health significance worldwide. The burden of disease caused by Cryptosporidium varies between and within countries/areas. To have a comprehensive understanding of epidemiological status and characteristics of human Cryptosporidium infection in China since the first report in 1987, a retrospective epidemiological analysis was conducted by presenting differences in the prevalence of Cryptosporidium by province, year, population, living environment and season and possible transmission routes and risk factors as well as genetic characteristics of Cryptosporidium in humans. METHODOLOGY/PRINCIPAL FINDINGS: A systematic search was conducted to obtain epidemiological papers of human Cryptosporidium infection/cryptosporidiosis from PubMed and Chinese databases. Finally, 164 papers were included in our analysis. At least 200,054 people from 27 provinces were involved in investigational studies of Cryptosporidium, with an average prevalence of 2.97%. The prevalence changed slightly over time. Variable prevalences were observed: 0.65-11.15% by province, 1.89-47.79% by population, 1.77-12.87% and 0-3.70% in rural and urban areas, respectively. The prevalence peak occurred in summer or autumn. Indirect person-to-person transmission was documented in one outbreak of cryptosporidiosis in a pediatric hospital. 263 Cryptosporidium isolates were obtained, and seven Cryptosporidium species were identified: C. hominis (48.3%), C. andersoni (22.43%), C. parvum (16.7%), C. meleagridis (8.36%), C. felis (3.04%), C. canis (0.76%) and C. suis (0.38%). CONCLUSIONS/SIGNIFICANCES: This systematic review reflects current epidemiological status and characteristics of Cryptosporidium in humans in China. These data will be helpful to develop efficient control strategies to intervene with and prevent occurrence of human Cryptosporidium infection/cryptosporidiosis in China as well as have a reference effect to other countries. Further studies should focus on addressing a high frequency of C. andersoni in humans and a new challenge with respect to cryptosporidiosis with an increasing population of elderly people and patients with immunosuppressive diseases.

First identification and genotyping of Enterocytozoon bieneusi in humans in Myanmar.

BACKGROUND: Intestinal pathogen infections are widespread among impoverished populations. Enterocytozoon bieneusi is the most common pathogen of intestinal microsporidian species in humans worldwide. However, no epidemiological information is available on E. bieneusi infection in humans in Myanmar. The present study comprised the first identification and genotyping of E. bieneusi in humans conducted in Myanmar. RESULTS: A total of 172 fecal specimens were collected from the Wa people (one each) in four villages of Pangsang Township of the Matman District of Shan State, Myanmar, and each participant completed a questionnaire. E. bieneusi was identified and genotyped using polymerase chain reaction (PCR) and sequence analysis of the internal transcribed spacer (ITS) region of the ribosomal RNA (rRNA) gene. The average prevalence of E. bieneusi was 8.72% (15/172), ranging from 3.85 to 13.89% by village. E. bieneusi infection was not related to any of the risk factors studied. Six genotypes were identified, comprising two known genotypes Peru6 (n = 10) and D (n = 1) and four novel genotypes (MMR23, MMR25, MMR86, and MMR87) (one each), and two people infected with genotype Peru6 were from the same family. A phylogenetic analysis based on a neighbor-joining tree of the ITS sequences of E. bieneusi indicated that all the six genotypes were clustered into group 1. CONCLUSIONS: This is the first identification and genotyping of E. bieneusi in humans in Myanmar. The observations that the two people infected with genotype Peru6 were from the same family, and that all six genotypes obtained in the present study fell into zoonotic group 1, showed the potential for anthropogenic and zoonotic transmissions. The present data argue for the importance of epidemiological control and prevention from medical sectors.

Identification and Evaluation of Reference Genes for Normalization of Gene Expression in Developmental Stages, Sexes, and Tissues of Diaphania caesalis (Lepidoptera, Pyralidae).

Diaphania caesalis (Walker) is an important boring insect mainly distributed in subtropical and tropical areas and attacked tropical woody grain crops, such as starchy plants of Artocarpus. Quantitative real-time polymerase chain reaction (qRT-PCR) is a powerful approach for investigating target genes expression profiles at the transcriptional level. However, the identification and selection of internal reference genes, which is often overlooked, is the most vital step before the analysis of target gene expression by qRT-PCR. So far, the reliable internal reference genes under a certain condition of D. caesalis have not been investigated. Therefore, this study evaluated the expression stability of eight candidate reference genes including ACT, ß-TUB, GAPDH, G6PDH, RPS3a, RPL13a, EF1α, and EIF4A in different developmental stages, tissues and sexes using geNorm, NormFinder and BestKeeper algorithms. To verify the stability of the recommended internal reference genes, the expression levels of DcaeOBP5 were analyzed under different treatment conditions. The results indicated that ACT, RPL13a, ß-TUB, RPS3a, and EF1α were identified as the most stable reference genes for further studies on target gene expression involving different developmental stages of D. caesalis. And ACT and EIF4A were recommended as stable reference genes for different tissues. Furthermore, ACT, EF1α, and RPS3a were ranked as the best reference genes in different sexes based on three algorithms. Our research represents the critical first step to normalize qRT-PCR data and ensure the accuracy of expression of target genes involved in phylogenetic and physiological mechanism at the transcriptional level in D. caesalia.

Validation of Asia-Pacific Colorectal Screening Score for Asymptomatic Participants and Outpatients in Northwest District of China.

INTRODUCTION: Asia-Pacific Colorectal Screening (APCS) score has been implemented for colorectal cancer screening in asymptomatic cohort in many regions. However, no study has validated its efficiency in Asian outpatients with mild-self-limited gastrointestinal symptoms yet. The purpose of this study was to validate its efficiency in asymptomatic subjects and outpatients in Ningxia. METHODS: The records of 329 asymptomatic participants and 300 outpatients were collected and analyzed from database in the General Hospital of Ningxia Medical University from September 2017 to April 2018. These 2 main groups were divided into 3 tiers based on the scores calculated by the category of APCS score. The detection rates of advanced colorectal neoplasia (ACRN) were further compared according to histopathological classifications of tissues acquired during colonoscopy. RESULTS: Among the 329 participants screened in the asymptomatic cohort, 78 subjects (23.7%) were in the low-risk (LR) tier, 187 subjects (56.8%) in the moderate-risk (MR) tier, and 64 subjects (19.5%) in the high-risk (HR) tier. The percentage of ACRN in the LR, MR, and HR groups was 1.3, 8.6, and 20.3%, respectively. In the 300 outpatient cohorts, 78 patients (26%) were in the LR tier, 140 patients (46.7%) in the MR tier, and 82 patients (27.3%) in the HR tier. The detection rates of ACRN in the LR, MR, and HR groups were 0, 10, and 39%, respectively. CONCLUSION: APCS score is an effective method for ACRN screening in asymptomatic and also the outpatient subjects. Individuals with HR scores should be given priority for colonoscopy.

Boronic acid sensors with double recognition sites: a review.

Boronic acids reversibly and covalently bind to Lewis bases and polyols, which facilitated the development of a large number of chemical sensors to recognize carbohydrates, catecholamines, ions, hydrogen peroxide, and so on. However, as the binding mechanism of boronic acids and analytes is not very clear, it is still a challenge to discover sensors with high affinity and selectivity. In this review, boronic acid sensors with two recognition sites, including diboronic acid sensors, and monoboronic acid sensors having another group or binding moiety, are summarized. Owing to double recognition sites working synergistically, the binding affinity and selectivity of sensors can be improved significantly. This review may help researchers to sort out the binding rules and develop ideal boronic acid-based sensors.

Evaluation and Validation of Reference Genes for Quantitative Real-Time PCR in Helopeltis theivora Waterhouse (Hemiptera: Miridae).

Helopeltis theivora Waterhouse is a predominant sucking pest in many tropic economic crops, such as tea, cocoa and coffee. Quantitative real-time PCR (qRT-PCR) is one of the most powerful tools to analyze the gene expression level and investigate the mechanism of insect physiology at transcriptional level. Gene expression studies utilizing qRT-PCR have been applied to numerous insects so far. However, no universal reference genes could be used for H. theivora. To obtain accurate and reliable normalized data in H. theivora, twelve candidate reference genes were examined under different tissues, developmental stages and sexes by using geNorm, NormFinder, BestKeeper, Delta Ct and RefFinder algorithms, respectively. The results revealed that the ideal reference genes differed across the treatments, and the consensus rankings generated from stability values provided by these programs suggested a combination of two genes for normalization. To be specific, RPS3A and Actin were the best suitable reference genes for tissues, RPL13A and GAPDH were suitable for developmental stages, EF1α and RPL13A were suitable for sexes, and RPL13A and RPS3A were suitable for all samples. This study represents the first systematic analysis of reference genes for qRT-PCR experiments in H. theivora, and the results can provide a credible normalization for qRT-PCR data, facilitating transcript profiling studies of functional genes in this insect.