This study investigated fecal specimens from 489 sheep and 537 cattle in multiple cities in northeast China for the prevalence and genetic characteristics of Enterocytozoon bieneusi by PCR and sequencing of the ribosomal internal transcribed spacer. Sixty-eight sheep specimens (13.9%) and 32 cattle specimens (6.0%) were positive for E. bieneusi. Sequence polymorphisms enabled the identification of 9 known genotypes (BEB4, BEB6, CM7, CS-4, EbpC, G, I, J, and OEB1) and 11 new genotypes (NESH1 to NESH6 and NECA1 to NECA5). The genotypes formed two genetic clusters in a phylogenetic analysis, with CS-4, EbpC, G, NESH1 to NESH3, and NECA1 to NECA5 distributed in zoonotic group 1 and BEB4, BEB6, CM7, EbpI, J, OEB1, and NESH4 to NESH6 distributed in potentially host-adapted group 2. Nearly 70% of cases of E. bieneusi infections in sheep were contributed by human-pathogenic genotypes BEB6, CS-4, and EbpC, and over 80% of those in cattle were by genotypes BEB4, CS-4, EbpC, I, and J. The cooccurrence of genotypes BEB4, CS-4, EbpC, I, and J in domestic ruminants and children in northeast China and the identification of BEB6 and EbpC in humans and water in central China imply the possibility of zoonotic transmission. This study also summarizes E. bieneusi genotypes obtained from ruminants worldwide and displays their host ranges, geographical distributions, and phylogenetic relationships. The data suggest a host range expansion in some group 2 genotypes (notably BEB4, BEB6, I, and J) that were previously considered to be adapted to ruminants. We should be concerned about the increasing zoonotic importance of group 2 genotypes with low host specificity. Microsporidia are a large and diverse group of obligately intracellular parasites that have been implicated as both human and animal pathogens (1). These parasitic protists are genetically related to fungi and feature environmentally resistant spore forms (1). Microsporidia differentiate from meronts into spores that are then defecated by the host into the environment and start a new round of eukaryotic cell invasion by using a highly specialized organelle, the polar tube, followed by intracellular replication (2). Of approximately 1,300 microsporidian species in 160 genera reported thus far, 14 species in 8 genera have been documented in human infections (3). Enterocytozoon bieneusi has emerged as an opportunistic pathogen leading to infectious diarrhea in humans; it has been associated with immune suppression and is responsible for almost 90% of reported cases of human microsporidiosis (4). It also affects immunocompetent individuals and a variety of domestic and wild animals, and even birds (4). Contact with infected humans and animals or contaminated food and water may contribute to the acquisition of E. bieneusi infections (1, 5-7).At present, genotyping of E. bieneusi on the basis of the ribosomal internal transcribed spacer (ITS) has characterized over 200 distinct genotypes (5). The genotype nomenclature used here is according to the established naming system (8). Cou...

As a kind of potent stimulus, lipopolysaccharide (LPS) has the ability to cause cell damage by activating toll-like receptor(TLR)4, then nuclear factor kappa B (NF-κB) translocates into the nucleus and changes the expression of related inflammatory genes. Baicalin is extracted from Radix Scutellariae, which possesses anti-inflammation, antioxidant and antibacterial properties. However, the effects of it on LPS-induced liver inflammation have not been fully elucidated. This study aims to investigate the anti-inflammatory effects of Baicalin on the LPS-induced liver inflammation and its underlying molecular mechanisms in chicken. The results of histopathological changes, serum biochemical analysis, NO levels and myeloperoxidase activity showed that Baicalin pretreatment ameliorated LPS-induced liver inflammation. ELISA and qPCR assays showed that Baicalin dose-dependently suppressed the production of IL-1β, IL-6, and TNF-α. Furthermore, the mRNA expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) were significantly decreased by Baicalin. TLR4 is an important sensor in LPS infection. Molecular studies showed that the expression of TLR4 was inhibited by Baicalin pretreatment. In addition, Baicalin pretreatment inhibited NF-kB signaling pathway activation. All results demonstrated the protective effects of Baicalin pretreatment against LPS-induced liver inflammation in chicken via negative regulation of inflammatory mediators through the down-regulation of TLR4 expression and the inhibition of NF-kB activation.

Enterocytozoon bieneusi is a well-known causative agent of microsporidial infections in a variety of mammal hosts including humans in China, whereas there were no epidemiological data on wild animals bred in captivity, and the role of the neglected hosts in transmission of zoonotic microsporidiasis remains unknown. Herein, we investigated feces from 191 farmed foxes (Vulpes vulpes) and 162 farmed raccoon dogs (Nyctereutes procyonoides) for the prevalence and genotypic characteristics of E. bieneusi in Harbin City, northeast China. Polymerase chain reaction (PCR) targeting the internal transcribed spacer (ITS) of the rRNA gene enabled the identification of 53 (27.7%) and 17 (10.5%) positives from fox and raccoon dog specimens, respectively. There was only minor difference in prevalence between juvenile and adult foxes. Adult raccoon dogs have an infection rate significantly higher than juveniles. The most common human-pathogenic E. bieneusi, genotype D, is widespread among foxes and raccoon dogs of various ages by sequence analysis of the ITS locus. Genotypes CHN-DC1 and mixed CHN-DC1/WildBoar3 were detected in one adult raccoon dog each. Here is the first report describing the presence of zoonotic E. bieneusi genotypes in farmed foxes and raccoon dogs. The widespread existence of genotype D in surveyed animals is of great concern for public health.

This study analyzed 563 fecal specimens of asymptomatic pigs from five cities of northeast China for the prevalence and genetic characteristics of Enterocytozoon bieneusi. The parasite was detected in 267 of 563 (47.4%) pigs by nested PCR of the ribosomal internal transcribed spacer (ITS). The differences in prevalence between preweaned (58.0%, 94/162) and growing pigs (39.6%, 114/288) and between weaned (52.2%, 59/113) and growing pigs are significant (p < 0.05). Genotypic typing and phylogenetic analysis facilitated identification of six human-pathogenic genotypes EbpC, O, CS-4, EbpA, Henan-IV, and PigEBITS5 and six potentially zoonotic genotypes EbpB, CC-1, CS-1, CS-3, CHN7, and CS-10. Genotypes CS-4 (32/35) and EbpC (3/35) from Harbin and Henan-IV (5/64) from Qiqihar determined in pigs herein represented the main causative agents of human microsporidiosis in Harbin. The most dominant genotype EbpC found in pigs from Daqing (35/65) and Qiqihar (a close neighbor to Daqing) (47/64) contributed significantly to human infections in Daqing. Genotype EbpC was also a leading E. bieneusi pathogen in humans, drinking water, and wastewater in central China. This study provided robust evidence that pigs could be an outstanding source of human microsporidiosis and water contamination in China.

It has been reported that oral intake of aflatoxin B1 ( AFB1 )-contaminated feed could cause acute, sub-chronic, or chronic toxicity in livestock and poultry. However, the harmful effect of AFB1 on the small intestine is still controversial. Therefore, blocking the entry of AFB1 into the body through the digestive tract is one of the important methods to prevent its toxicity. In the present study, 1-day-old Arbor Acres broilers were randomly divided into 6 groups including control group, curcumin control group (450 mg curcumin/kg feed), curcumin low-, medium-, and high-dose group (150, 300, and 450 mg curcumin/kg feed + 5 mg AFB1/kg feed), and AFB1 group (5 mg AFB1/kg feed). After 28 d, the samples of chickens' duodenums were collected for further analyses. AFB1 caused abnormal functional and morphological changes in the duodenum, including histological lesions, increased the length of the duodenum and depth of crypt, decreased the unit weight of the duodenum, height of villus, and the value of villus height/crypt depth. Meanwhile, AFB1 administration enhanced malonaldehyde activity, 8-HOdG level, and the mRNA expression of cytochrome P450 ( CYP450 ) enzymes, and reduced superoxide dismutase, catalase, adenosine triphosphatase ( ATPase ) activity and the mRNA expression of Abcb1 . Importantly, curcumin supplementation partially ameliorated AFB1-induced abnormal functional and morphological signs of the duodenum, alleviated AFB1-induced oxidative stress, and decreased the mRNA expression of CYP450 enzymes. Furthermore, curcumin ameliorated AFB1-induced decrease in the Abcb1 mRNA expression, P-glycoprotein ( P-gp ) level, and ATPase activities. It has been suggested from these results that curcumin supplementation in the feed could ameliorate AFB1-induced duodenal toxicity and damage through downregulating CYP450 enzymes, promoting ATPase activities, and inducing P-gp in chickens.

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BackgroundImproper use of antimicrobials results in poor treatment and severe bacterial resistance. Breakpoints are routinely used in the clinical laboratory setting to guide clinical decision making. Therefore, the objective of this study was to establish antimicrobial susceptibility breakpoints for danofloxacin against Escherichia coli (E.coli), which is an important pathogen of digestive tract infections.ResultsThe minimum inhibitory concentrations (MICs) of 1233 E. coli isolates were determined by the microdilution broth method in accordance with the guidelines in Clinical and Laboratory Standards Institute (CLSI) document M07-A9. The wild type (WT) distribution or epidemiologic cutoff value (ECV) was set at 8 μg/mL with statistical analysis. Plasma drug concentration data were used to establish pharmacokinetic (PK) model in swine. The in vitro time kill test in our study demonstrated that danofloxacin have concentration dependent activity against E.coli. The PK data indicated that danofloxacin concentration in plasma was rapidly increased to peak levels at 0.97 h and remained detectable until 48 h after drug administration. The pharmacodynamic cutoff (COPD) was determined as 0.03 μg/mL using Monte Carlo simulation. To the best of our knowledge, this is the first study to establish the ECV and COPD of danofloxacin against E.coli with statistical method.ConclusionsCompared to the COPD of danofloxacin against E.coli (0.03 μg/mL), the ECV for E.coli seemed reasonable to be used as the final breakpoint of danofloxacin against E.coli in pigs. Therefore, the ECV (MIC ≤8 μg/mL) was finally selected as the optimum danofloxacin susceptibility breakpoint for swine E.coli. In summary, this study provides a criterion for susceptibility testing and improves prudent use of danofloxacin for protecting public health.

Aflatoxin B1 (AFB1) and its metabolite aflatoxin M1 (AFM1) are well-known carcinogens for humans and animals health. In this study, an ultra-high performance liquid chromatography linked with fluorescence detection (UPLC-FLD) method was optimized and validated. In addition, we investigated for the first time, the influence of curcumin on residue depletion of AFB1 and AFM1 in liver, kidney, and muscle tissues of broiler chickens and estimated a necessary clearance time required for AFB1 and AFM1 residues. The results showed that the average recoveries of AFB1 varied in liver, kidney, and muscles between 82.32–85.56, 85.34–88.45, and 84.88–89.73% respectively, while the average recoveries of AFM1 in liver, kidney, and muscles varied between 92.17–95.03, 94.12–97.21, and 95.32–98.51%, respectively. The detection limit of aflatoxin B1 was 0.008 ng/ml, while for aflatoxin M1 was 0.003 ng/ml. The limit of quantification (LOQ) for AFB1 and AFM1 was 0.02 and 0.01 ng/ml, respectively. Clearance time for AFB1 and AFM1 residues were analyzed in two experimental groups of broilers. One group fed with dietary AFB1 (5.0 mg/kg feed) and other with curcumin+AFB1 diet (curcumin; 300 mg/kg feed, AFB1; 5.0 mg/kg feed). AFB1 and AFM1 residues clearance time was calculated based on LOQ using withdrawal time calculation software (WT1.4). Clearance time analyzed for AFB1 ranged from 11 to 19 days and for AFM1 ranged from 10 to 12 days at 95% confidence level. Interestingly, curcumin supplementation in the diet reduced the clearance time of AFM1 in liver and kidney but not in muscle tissues. Conclusively, the developed method can be appropriately used for the quality control testing of commercial broiler-meat processing companies, food manufacturers, and quality control laboratories.