Epidemiological investigation of coccidiosis and associated risk factors in broiler chickens immunized with live anticoccidial vaccines in China.

Coccidiosis is a costly intestinal disease of chickens caused by Eimeria species. This infection is associated with high mortality, reduced feed efficiency, and slowed body weight gain. The diagnosis and control of coccidiosis becomes challenging due to the fact that chickens can be infected by seven different Eimeria species and often occur mixed-species co-infections. Grasping the epidemiology of Eimeria species is crucial to estimate the efficiency of poultry management. This study aimed to explore the distribution of Eimeria species in broiler chickens in China after administering live anticoccidial vaccines. A total of 634 samples were obtained, and the survey results showed that the prevalence of Eimeria was 86.12% (546/634), and the most common species were E. acervulina (65.62%), E. necatrix (50.95%), E. mitis (50.79%), E. tenella (48.42%), and E. praecox (41.80%). Most samples indicated mixed-species infections (an average of 3.29 species per positive sample). Notably, 63.98% of samples contain 3 to 5 Eimeria species within a single fecal sample. The most prevalent combinations were E. acervulina-E. tenella (38.96%) and E. acervulina-E. necatrix (37.22%). Statistical analysis showed that flocks vaccinated with trivalent vaccines were significantly positive for E. necatrix in grower chickens (OR = 3.30, p < 0.05) compared with starter chickens, and tetravalent vaccinated flocks showed that starter chickens demonstrated a higher susceptibility to E. tenella-E. brunetti (OR = 2.03, p < 0.05) and E. acervulina-E. maxima (OR = 2.05, p < 0.05) compared with adult chickens. Geographically, in the case of tetravalent vaccine-immunized flocks, a substantial positive association was observed between E. necatrix infection rates and flocks from eastern (OR = 3.88, p < 0.001), central (OR = 2.65, p = 0.001), and southern China (OR = 3.17, p < 0.001) compared with southwestern China. This study also found a positive association between E. necatrix (OR = 1.64, p < 0.05), E. acervulina (OR = 1.59, p < 0.05), and E. praecox (OR = 1.81, p < 0.05) infection and coccidiosis occurrence compared with non-infected flocks in tetravalent vaccinated flocks. This molecular epidemiological investigation showed a high prevalence of Eimeria species in the field. The emergent species, E. brunetti and E. praecox, might be incorporated into the widely-used live vaccines in the future. These insights could be useful in refining coccidiosis control strategies in the poultry industry.

Genotypic diversity and epidemiology of Trichomonas gallinae in Columbidae: Insights from a comprehensive analysis.

Trichomonas gallinae is a protozoa that parasitizes the upper gastrointestinal and respiratory tracts of various animals and birds, including Columbidae, Passeriformes, and Falconiformes. Polymerase chain reaction-based T. gallinae ITS1/5.8S/ITS2 gene typing yields inconsistent results owing to methodological differences. To standardize the statistical analysis of T. gallinae genotype distributions, this study employed MEGA-X software with the Tamamura 3-parameter (T92) + G model in the neighbor-joining method, with 2,000 bootstrap replicates, to calculate a systematic evolutionary tree. The resulting tree comprised 12 branches, ITS-OBT-Tg-1 to ITS-OBT-Tgl, with similar phylogenetic relationships. Relevant literature review yielded T. gallinae prevalence data in Columbidae. Statistical analysis was conducted from two perspectives: non-biological and biological factors, using chi-square tests and ordered logistic regression analysis. T. gallinae positivity rates differed significantly across diverse regions (χ2 = 4,609.9, P = 0.000, df = 4) and at various times (χ2 = 2,810.8, P = 0.000, df = 3). However, temperature and precipitation did not significantly affect T. gallinae positivity rates. Additionally, T. gallinae positivity rates differed significantly among diverse hosts (χ2 = 2,958.6, P = 0.000, df = 14) and by host age (χ2 = 478.5, P = 0.000, df = 2) and sex (χ2 = 96.00, P = 0.000, df = 1). This comprehensive analysis aimed to control T. gallinae transmission, reduce economic and species resource losses, and provide a foundation for future related research.

Unraveling the pathogenic potential of the Pentatrichomonas hominis PHGD strain: impact on IPEC-J2 cell growth, adhesion, and gene expression.

Pentatrichomonas hominis, a flagellated parasitic protozoan, predominantly infects the mammalian digestive tract, often causing symptoms such as abdominal pain and diarrhea. However, studies investigating its pathogenicity are limited, and the mechanisms underlying P. hominis-induced diarrhea remain unclear. Establishing an in vitro cell model for P. hominis infection is imperative. This study investigated the interaction between P. hominis and IPEC-J2 cells and its impact on parasite growth, adhesion, morphology, and cell viability. Co-cultivation of P. hominis with IPEC-J2 cells resulted in exponential growth of the parasite, with peak densities reaching approximately 4.8 × 105 cells/mL and 1.2 × 106 cells/mL at 48 h for initial inoculation concentrations of 104 cells/mL and 105 cells/mL, respectively. The adhesion rate of P. hominis to IPEC-J2 cells reached a maximum of 93.82% and 86.57% at 24 h for initial inoculation concentrations of 104 cells/mL and 105 cells/mL, respectively. Morphological changes in IPEC-J2 cells co-cultivated with P. hominis were observed, manifesting as elongated and irregular shapes. The viability of IPEC-J2 cells exhibited a decreasing trend with increasing P. hominis concentration and co-cultivation time. Additionally, the mRNA expression levels of IL-6, IL-8, and TNF-α were upregulated, whereas those of CAT and CuZn-SOD were downregulated. These findings provide quantitative evidence that P. hominis can promote its growth by adhering to IPEC-J2 cells, inducing morphological changes, reducing cell viability, and triggering inflammatory responses. Further in vivo studies are warranted to confirm these results and enhance our understanding of P. hominis infection.

Title: Découvrir le potentiel pathogène de la souche PHGD de Pentatrichomonas hominis : impact sur la croissance, l'adhésion et l'expression des gènes des cellules IPEC-J2. Abstract: Pentatrichomonas hominis, un protozoaire parasite flagellé, infecte principalement le tube digestif des mammifères, provoquant souvent des symptômes tels que des douleurs abdominales et de la diarrhée. Cependant, les études portant sur sa pathogénicité sont limitées et les mécanismes sous-jacents à la diarrhée induite par P. hominis restent flous. L'établissement d'un modèle cellulaire in vitro de l'infection à P. hominis est impératif. Cette étude a examiné l'interaction entre P. hominis et les cellules IPEC-J2 et son impact sur la croissance du parasite, l'adhésion, la morphologie et la viabilité cellulaire. La co-culture de P. hominis avec des cellules IPEC-J2 a entraîné une croissance exponentielle du parasite, avec des densités maximales atteignant environ 4,8 × 105 cellules/mL et 1,2 × 106 cellules/mL à 48 h pour des concentrations d'inoculation initiales de 104 cellules/mL et 105 cellules/mL, respectivement. Le taux d'adhésion de P. hominis aux cellules IPEC-J2 a atteint un maximum de 93,82 % et 86,57 % après 24 h pour des concentrations d'inoculation initiales de 104 cellules/mL et 105 cellules/mL, respectivement. Des changements morphologiques dans les cellules IPEC-J2 co-cultivées avec P. hominis ont été observés, se manifestant par des formes allongées et irrégulières. La viabilité des cellules IPEC-J2 a montré une tendance à la baisse avec l'augmentation de la concentration de P. hominis et de la durée de co-culture. De plus, les niveaux d'expression d'ARNm d'IL-6, d'IL-8 et de TNF-α étaient régulés positivement, tandis que ceux de CAT et de CuZn-SOD étaient régulés négativement. Ces résultats fournissent des preuves quantitatives que P. hominis peut favoriser sa croissance en adhérant aux cellules IPEC-J2, en induisant des changements morphologiques, en réduisant la viabilité cellulaire et en déclenchant des réponses inflammatoires. D'autres études in vivo sont nécessaires pour confirmer ces résultats et améliorer notre compréhension de l'infection à P. hominis.

Immunoprotective effects of DNA vaccine against Eimeria tenella based on EtAMA3 and EtRON2L2.

Eimeria tenella is the most pathogenic and harmful intestinal parasitic protozoan. Recombinant DNA vaccines open options for promising strategies for preventing avian coccidiosis, replacing chemical drugs and live oocyst vaccines. Two important antigenic proteins, EtAMA3 (also known as SporoAMA1) and EtRON2L2, act together to promote the invasion of E. tenella sporozoites. In this study, a recombinant DNA vaccine, designated pcDNA3.1(+)-AR, was constructed based on EtAMA3DII, EtRON2L2D3, and EtRON2L2D4. Chickens were intramuscularly immunized with different doses (25, 50, or 100 µg) of pcDNA3.1(+)-AR to evaluate its immunoprotective effects in vivo. The chickens in the 50 µg and 100 µg groups had higher cytokine concentrations (interleukin 2, interferon-gamma, and interleukin 10), and lesion scores (81.9% and 67.57%, respectively) and relative oocyst production (47% and 19%, respectively) reduced compared with the unchallenged group, indicating partial protection against E. tenella. These results suggest that pcDNA3.1(+)-AR is a promising vaccine candidate against avian coccidiosis.

Drug resistance patterns and genotype associations of Trichomonas gallinae in meat pigeons (Columba livia): insights from Guangdong Province, China.

Avian trichomoniasis, caused by the protozoan parasite Trichomonas gallinae, is a prevalent and economically significant disease in pigeons. This study investigated the drug resistance of T. gallinae isolates in Guangdong Province, China. The results revealed that 25.3% (20/79) of the isolates were resistant to one or more of the four nitroimidazole drugs tested, namely, metronidazole, dimetridazole, secnidazole, and tinidazole. Secnidazole elicited the highest resistance rate (19.0%; 15/79), followed by tinidazole (17.7%; 14/79), metronidazole (17.7%; 14/79), and dimetridazole (13.9%; 11/79). An enormous majority of the resistant isolates (70.0%; 14/20) exhibited resistance to multiple drugs. Additionally, the resistance rate was significantly higher in isolates from birds aged < 30 days (53.3%; 8/15) than in those from older birds (23.1%; 12/52). Moreover, no drug resistance was detected in female pigeons. The genotype of the isolated strain was also associated with drug resistance. Specifically, 50.0% (15/30) of ITS-B genotypes exhibited resistance to drugs, while only 10.2% (5/49) of ITS-A genotypes demonstrated resistance. This study also found the growth characteristics of different Trichomonas isolates to be influenced by their genotypes and initial inoculum concentrations. These findings underscore the urgent need for effective measures to control and prevent drug-resistant T. gallinae infections in pigeons, thus ensuring the stable development of the pigeon industry.

One-Step Synergistic Treatment Approach for High Performance Amorphous InGaZnO Thin-Film Transistors Fabricated at Room Temperature.

Amorphous InGaZnO (a-InGaZnO) is currently the most prominent oxide semiconductor complement to low-temperature polysilicon for thin-film transistor (TFT) applications in next-generation displays. However, balancing the transmission performance and low-temperature deposition is the primary obstacle in the application of a-InGaZnO TFTs in the field of ultra-high resolution optoelectronic display. Here, we report that a-InGaZnO:O TFT prepared at room temperature has high transport performance, manipulating oxygen vacancy (VO) defects through an oxygen-doped a-InGaZnO framework. The main electrical properties of a-InGaZnO:O TFTs included high field-effect mobility (µFE) of 28 cm2/V s, a threshold voltage (Vth) of 0.9 V, a subthreshold swing (SS) of 0.9 V/dec, and a current switching ratio (Ion/Ioff) of 107; significant improvements over a-InGaZnO TFTs without oxygen plasma. A possible reason for this is that appropriate oxygen plasma treatment and room temperature preparation technology jointly play a role in improving the electrical performance of a-InGaZnO TFTs, which could not only increase carrier concentration, but also reduce the channel-layer surface defects and interface trap density of a-InGaZnO TFTs. These provides a powerful way to synergistically boost the transport performance of oxide TFTs fabricated at room temperature.

Essential Functions of Calmodulin and Identification of Its Proximal Interacting Proteins in Tachyzoite-Stage Toxoplasma gondii via BioID Technology.

Toxoplasma gondii (T. gondii) is a pathogen belonging to the apicomplexan phylum, and it threatens human and animal health. Calcium ions, a critical second messenger in cells, can regulate important biological processes, including parasite invasion and egress. Calmodulin (CaM) is a small, highly conserved, Ca2+-binding protein found in all eukaryotic cells. After binding to Ca2+, CaM can be activated to interact with various proteins. However, little is known about CaM's function and its interacting proteins in T. gondii. In this study, we successfully knocked down CaM in the T. gondii parent strain TATI using a tetracycline-off system with the Toxoplasma CaM promoter. The results indicated that CaM was required for tachyzoite proliferation, invasion, and egress and that CaM depletion resulted in apicoplast loss, thus threatening parasite survival in the next lytic cycle. In the tachyzoite stage, CaM loss caused significant anomalies in the parasite's basal constriction, motility, and parasite rosette-like arrangement in the parasitophorous vacuole (PV). These phenotypic defects caused by CaM depletion indicate the importance of CaM in T. gondii. Therefore, it is important to identify the CaM-interacting proteins in T. gondii. Applying BioID technology, more than 300 CaM's proximal interacting proteins were identified from T. gondii. These CaM partners were broadly distributed throughout the parasite. Furthermore, the protein interactome and transcriptome analyses indicated the potential role of CaM in ion binding, cation binding, metal ion binding, calcium ion binding, and oxidation-reduction. Our findings shed light on the CaM function and CaM-interactome in T. gondii and other eukaryotes. IMPORTANCE Toxoplasma gondii is an intracellular pathogen that threatens human and animal health. This unicellular parasite is active in many biological processes, such as egress and invasion. The implementation efficiency of T. gondii biological processes is dependent on signal transmission. Ca2+, as a second messenger, is essential for the parasite's life cycle. Calmodulin, a ubiquitous Ca2+ receptor protein, is highly conserved and mediates numerous Ca2+-dependent events in eukaryotes. Few CaM functions or regulated partners have been characterized in T. gondii tachyzoites. Here, we reported the essential functions of calmodulin in T. gondii tachyzoite and the identification of its interacting partners using BioID technology, shedding light on the CaM function and CaM-interactome in Toxoplasma gondii and other eukaryotes.

Evolution Application of Two-Dimensional MoS2-Based Field-Effect Transistors.

High-performance and low-power field-effect transistors (FETs) are the basis of integrated circuit fields, which undoubtedly require researchers to find better film channel layer materials and improve device structure technology. MoS2 has recently shown a special two-dimensional (2D) structure and superior photoelectric performance, and it has shown new potential for next-generation electronics. However, the natural atomic layer thickness and large specific surface area of MoS2 make the contact interface and dielectric interface have a great influence on the performance of MoS2 FET. Thus, we focus on its main performance improvement strategies, including optimizing the contact behavior, regulating the conductive channel, and rationalizing the dielectric layer. On this basis, we summarize the applications of 2D MoS2 FETs in key and emerging fields, specifically involving logic, RF circuits, optoelectronic devices, biosensors, piezoelectric devices, and synaptic transistors. As a whole, we discuss the state-of-the-art, key merits, and limitations of each of these 2D MoS2-based FET systems, and prospects in the future.

Characterization and evaluation of a recombinant multiepitope peptide antigen MAG in the serological diagnosis of Toxoplasma gondii infection in pigs.

BACKGROUND: Toxoplasmosis caused by Toxoplasma gondii is a serious disease threatening human and animal health. People can be infected with T. gondii by ingesting raw pork contaminated with cysts or oocysts. Serological test is a sensitive and specific method usually used for large-scale diagnosis of T. gondii infection in humans and animals (such as pigs). Commercial pig Toxoplasma antibody ELISA diagnostic kits are expensive, which limits their use; moreover, the wide antigen composition used in these diagnostic kits is still unclear and difficult to standardize. The multiepitope peptide antigen is a novel diagnostic marker, and it has potential to be developed into more accurate and inexpensive diagnostic kits. METHODS: The synthetic multiepitope antigen (MAG) cDNA encoding a protein with epitopes from five T. gondii-dominant antigens (SAG1, GRA1, ROP2, GRA4, and MIC3) was designed, synthesized, and expressed in Escherichia coli BL21 (DE3) strain. The recombinant protein was detected through western blot with pig anti-T. gondii-positive and -negative serum, and then IgG enzyme-linked immunosorbent assay (ELISA) named MAG-ELISA was designed. The MAG-ELISA was evaluated in terms of specificity, sensitivity, and stability. The MAG-ELISA was also compared with a commercial PrioCHECK® Toxoplasma Ab porcine ELISA (PrioCHECK ELISA). Finally, the trend of pig anti-T. gondii IgG levels after artificial infection with RH tachyzoites was evaluated using MAG-ELISA and two other ELISA methods (rMIC3-ELISA and PrioCHECK ELISA). RESULTS: MAG antigen could be specifically recognized by pig anti-T. gondii-positive but not -negative serum. MAG-ELISA showed high diagnostic performance in terms of specificity (88.6%) and sensitivity (79.1%). MAG-ELISA could be used for detecting anti-T. gondii IgG in the early stage of T. gondii infection in pigs (at least 7 days after artificial infection). CONCLUSIONS: Our results suggest that MAG antigen can be applied to specifically recognize anti-T. gondii IgG in pig, and MAG-ELISA has the potential for large-scale screening tests of T. gondii infection in pig farms and intensive industries.

A Comparison of Transcriptional Diversity of Swine Macrophages Infected With TgHB1 Strain of Toxoplasma gondii Isolated in China.

Toxoplasma gondii is an apicomplexan parasite infecting human and animals, causing huge public health concerns and economic losses. Swine alveolar macrophage plays an important role in controlling T. gondii infection. However, the mechanism by which macrophages infected with T. gondii function in the immunity to the infection is unclear, especially for local isolates such as TgHB1 isolated in China. RNA-seq as a valuable tool was applied to simultaneously analyze transcriptional changes of pig alveolar macrophages infected with TgRH (typeI), TgME49 (typeII) or TgHB1 at different time points post infection (6, 12, and 24 h). Paired-end clean reads were aligned to the Sscrofa10.2 pig genome and T. gondii ME49 genome. The differentially expressed genes of macrophages and T. gondii were enriched through Gene Ontology and Kyoto Encyclopedia of Genes and Genomes, respectively. Compared to the TgRH and TgME49 infection groups, 307 down-regulated macrophage genes (mainly enriched for development and metabolism) and 419 up-regulated genes (mainly enriched for immune pathways) were uniquely expressed in the TgHB1 infection group. Additionally, 557 down-regulated and 674 up-regulated T. gondii genes (mainly enriched in metabolism and biosynthesis) were uniquely expressed in the TgHB1 infection group. For validation purposes, some of the differentially expressed genes of macrophages involved in immune-related signaling pathways were used for further analysis via real time quantitative reverse-transcription polymerase-chain reaction (qRT-PCR). This work provides important insights into the temporal immune responses of swine alveolar macrophages to infection by the strain TgHB1 isolated from China, and is helpful for better understanding of the T. gondii genotype-associated activation of macrophages during early phase of the infection.

Identification of Novel Dense-Granule Proteins in Toxoplasma gondii by Two Proximity-Based Biotinylation Approaches.

Toxoplasma gondii is an opportunistic pathogen infecting humans and a variety of vertebrate animals. Secretory dense-granule proteins (GRAs) play diverse roles in the mediation of host-parasite interactions and facilitate parasitism, but many of them still remain to be identified. Here, we used two proximity-based protein labeling techniques to identify novel GRA proteins. Taking GRA1 as bait, transgenic strains expressing GRA1-BirA* or GRA1-APEX were constructed to biotinylate GRAs. Using these methods, a total of 46 proteins were identified, 20 of which were known GRA proteins. Among these 46, 17 were identified by both strategies, and 14 out of the 17 were known GRAs. The other three were all confirmed to localize to dense granules. Nonetheless a significant portion of the proteins were only identified by either APEX or BirA*, indicating that there are differences between these methods. Of the 26 novel GRAs, 5 were validated as bona fide GRAs by localization studies. The majority of these novel GRAs are only present in coccidian parasites and are likely dispensable for parasite growth in vitro; they may play roles during animal infections. The identification of novel GRAs laid the foundation for further studies investigating the mechanisms underlying parasite-host interactions.

Screening and Identification of the Host Proteins Interacting with Toxoplasma gondii Rhoptry Protein ROP16.

Toxoplasma gondii, as a zoonotic protozoan parasite, develops sophisticated strategies to manipulate hosts for efficient intracellular survival. After successful invasion, T. gondii injects many effector proteins into host cells for various purposes. TgROP16 (T. gondii rhoptry protein 16), which is secreted from rhoptries into host cells, can activate the host STAT (signal transducer and activator of transcription) signaling pathway through phosphorylation of STAT3 and STAT6. However, whether there are other host proteins modulated by TgROP16 is currently unknown. In this study, yeast two-hybrid (Y2H) screen was used to look for additional host proteins interacting with TgROP16. Yeast cells expressing a mouse cDNA library cloned into the prey vector were used to mate with yeasts expressing ROP16 without signal peptide. Two mouse proteins, Dnaja1 (DnaJ heat shock protein family member A1) and Gabra4 (gamma-aminobutyric acid A receptor, subunit alpha 4) were identified to interact with ROP16 from this screen. Further analysis suggested that the Predomain of ROP16 played key roles in mediating interactions with these host proteins, whereas the contribution from the Kinase domain was minor. The interactions between Dnaja1 and different parts of ROP16 were also estimated in vivo by co-immunoprecipitation. The results showed that the Predomain of ROP16 was the major region to interact with Dnaja1, which is consistent with the Y2H results. Based on the gene ontology analysis, Dnaja1 is predicted to participate in stress response while Gabra4 is involved in the system development process. The discovery of new host proteins that interact with ROP16 of T. gondii will help us to further investigate the functions of this effector proteins during T. gondii infection.