Microscopic extrathyroidal extension does not affect the prognosis of patients with papillary thyroid carcinoma: A propensity score matching analysis.

Background: Extrathyroidal extension (ETE) in papillary thyroid carcinoma (PTC) can be divided into two categories based on different degrees of invasion: microscopic ETE (micro-ETE) and macroscopic ETE (macro-ETE). At present, there is a consensus that macro-ETE significantly affects PTC prognosis, while the prognostic significance of micro-ETE remains controversial. Methods: The clinicopathological and follow-up data for PTC patients who underwent surgical treatment at the Hangzhou First People's Hospital between 2015 and 2018 were retrospectively analyzed. According to the degree of ETE, patients were divided into three groups: non-ETE, micro-ETE and macro-ETE. Cox regression analysis was performed to evaluate the effect of ETE on recurrence-free survival (RFS). The propensity score matching (PSM) method was used to reduce the interference of confounding factors, and Kaplan-Meier curves were utilized to compare the RFS. Results: Both micro- and macro-ETE were associated with some aggressive tumor features, including tumor size, multifocality, and lymph node metastasis. Univariate and multivariate Cox regression analyses showed that macro-ETE was an independent risk factor for recurrence, while micro-ETE was not associated with recurrence. The K-M curves showed that RFS for micro-ETE and non-ETE were not statistically different before and after PSM, while RFS for macro-ETE was significantly shorter than that for non-ETE. Conclusion: The presence of micro-ETE in PTC did not affect prognosis of patients, suggesting that its treatment should be consistent with the treatment for intrathyroidal tumors. The surgical method and the necessity for radioiodine therapy should be carefully evaluated to reduce overtreatment.

First Molecular Detection and Genotype Identification of Toxoplasma gondii in Chickens from Farmers' Markets in Fujian Province, Southeastern China.

Toxoplasma gondii is an opportunistic pathogenic protozoan that can infect all nucleated cells in almost all warm-blooded animals, including humans. T. gondii infection has been reported in many food animals worldwide. However, the prevalence and genotypes of T. gondii in chickens from farmers' markets in Fujian province in southeastern China remain unreported. In the present study, four tissue samples from each of the 577 chickens (namely, the heart, liver, lungs, and muscles) were collected from farmers' markets in five regions of Fujian province (Zhangzhou, Sanming, Quanzhou, Fuzhou, and Longyan). We first analyzed the prevalence and genotypes of T. gondii using PCR targeting of the B1 gene of T. gondii. Of the 577 chickens, thirty-two (5.5%) tested positive for the B1 gene. Among the five regions, Sanming had the highest infection rate (16.8%, 16/95), followed by Quanzhou (8.0%, 8/100), Longyan (5.0%, 5/100), Zhangzhou (1.1%, 2/182), and Fuzhou (1.0%, 1/100). Among these thirty-two T. gondii-positive chickens, the infection rates of the lungs, heart, liver, and muscles were 68.8% (22/32), 34.4% (11/32), 28.1% (9/32), and 9.4% (3/32), respectively. Significant differences in prevalence were found among the different regions (χ2 = 35.164, p < 0.05) and tissues (χ2 = 25.874, p < 0.05). A total of 128 tissue and organ samples of the thirty-two T. gondii-positive chickens from the different regions were analyzed using PCR-restriction fragment length polymorphism (PCR-RFLP) on the basis of 10 genetic markers. Seven tissue samples (lung samples from five chickens, heart samples from one chicken, and liver samples from one chicken) underwent successful amplification at all the genetic markers, and all the T. gondii genotypes were identified as genotype I (ToxoDB #10). These findings serve as a foundation for evaluating the risk of T. gondii contamination in chicken products intended for human consumption and offer insight into preventing the transmission of the parasite from chickens to humans.

Proteomics analyses of acute kidney injury biomarkers in a rat exertional heat stroke model.

The frequency of exertional heat stroke (EHS) increases with the gradual elevation of global temperatures during summer. Acute kidney injury (AKI) is a common complication of EHS, and its occurrence often indicates the worsening of a patient's condition or a poor prognosis. In this study, a rat model of AKI caused by EHS was established, and the reliability of the model was evaluated by HE staining and biochemical assays. The expression of kidney tissue proteins in the EHS rats was analyzed using label-free liquid chromatography-tandem mass spectrometry. A total of 3,129 differentially expressed proteins (DEPs) were obtained, and 10 key proteins were finally identified, which included three upregulated proteins (Ahsg, Bpgm, and Litaf) and seven downregulated proteins (medium-chain acyl-CoA synthetase 2 (Acsm2), Hadha, Keg1, Sh3glb1, Eif3d, Ambp, and Ddah2). The qPCR technique was used to validate these 10 potential biomarkers in rat kidney and urine. In addition, Acsm2 and Ahsg were double-validated by Western blotting. Overall, this study identified 10 reliable biomarkers that may provide potential targets for the treatment of AKI caused by EHS.

Heat shock protein 60 in parasitic helminths: A role in immune responses and therapeutic applications.

Heat shock protein 60 (HSP60) is an unique member of the heat shock protein family, being involved in parasite infections. To cope with harsh environments where parasites live, HSP60s are indispensable and involved in a variety of biological processes. HSP60s have relative low similarity among parasites, but their ATPase /Mg2+ active sites are highly conserved. The interactions of HSP60s with signaling pathway regulators in immune cells suggest a crucial role in immune responses, rendering them a potential therapeutic target. This paper reviews the current understandings of HSP60s in parasitic helminths in aspects of molecular characteristics, immunoregulatory responses and HSP60-based therapeutics.

Comprehensive analysis of mRNA-lncRNA co-expression profiles in mouse brain during infection with Toxoplasma gondii.

Toxoplasma gondii is an obligate intracellular protozoan parasite which seriously threatens the health of domestic animals and humans. Long non-coding RNAs (lncRNAs) are non-protein-coding transcripts greater than 200 nucleotides, which are widely involved in transcriptional and epigenetic regulations. However, little is known about the roles of host lncRNAs in the response to T. gondii infections. In this study, using Illumina sequencing technology, we analyzed the expression profiles of mRNAs and lncRNAs in BALB/c mouse brain following infection by T. gondii PRU strain (type II genotype) cysts. The identified differentially expressed (DE) RNAs were subjected to bioinformatics analysis. A total of 2,090 annotated lncRNAs along with 3,577 novel lncRNAs were identified. In the acutely infected mouse brain, a total of 330 mRNAs and 19 lncRNAs were dys-regulated, whereas 136 DE mRNAs and 9 DE lncRNAs were identified in chronically infected mouse brain. GO analysis revealed that these DE mRNAs identified at acute infection stage were involved in immune response, whereas DE mRNAs found at chronic infection stage were mostly enriched in response to protozoan. KEGG analysis showed that DE mRNAs were significantly enriched in disease related pathways. In addition, the putative mRNA-lncRNA co-expression network was constructed, and several hub regulatory RNAs were identified based on the transcriptome data. This study firstly characterized the co-expression profile of mRNAs and lncRNAs in mouse brain infected with T. gondii and provided a framework for further studies of the roles of lncRNAs in host neuropathology during toxoplasmosis progression.

Molecular Detection and Multilocus Genotyping of Giardia duodenalis in Pigs in Fujian Province, Southeastern China.

Giardia duodenalis, an intestinal parasite, is widely distributed in humans and various animals, such as pigs, cattle and cats. The clinical symptoms of giardiasis are characterized as including abdominal pain, acute or chronic diarrhea, and bloating and weight loss in humans and animals, leading to public and veterinary health problems worldwide. However, the prevalence and genotypes of G. duodenalis in pigs in Fujian Province, southeastern China, have not been reported. In the present study, 725 fecal samples were collected from six cities (Fuqing, Putian, Nanping, Longyan, Sanming, Zhangzhou) in Fujian Province and analyzed for G. duodenalis prevalence and genotypes using nested PCR targeting the beta-giardin (bg), glutamate dehydrogenase (gdh) and triosephosphate isomerase (tpi) genes. The results shown that total occurrence rate of G. duodenalis was 26.9% (195/725) in pigs, with significant differences in the prevalence among different regions (χ2 = 86.508, p < 0.05) and groups (χ2 = 12.748, p < 0.05). 195, 11 and 6 samples were detected at the bg, tpi and gdh loci, respectively. Each one belonged to a subtype of assemblage E and was analyzed using sequences obtained in this study. Based on phylogenetic analyses of sequences from the three genetic loci, only one MLG E1 was found. The results indicated that pigs may present a potential zoonotic risk of spreading G. duodenalis infection from animals to humans in this area. The findings of the present study also provide basic data for the prevention and control of G. duodenalis infection in pigs and humans in China.

Urine proteomics for profiling of mouse toxoplasmosis using liquid chromatography tandem mass spectrometry analysis.

BACKGROUND: Toxoplasma gondii is an obligate intracellular parasite that causes toxoplasmosis. Urine is an easily obtained clinical sample that has been widely applied for diagnostic purposes. However, changes in the urinary proteome during T. gondii infection have never been investigated. METHODS: Twenty four-hour urine samples were obtained from BALB/c mice with acute infection [11 days post infection (DPI)], mice with chronic infection (35 DPI) and healthy controls, and were analyzed using a label-free liquid chromatography tandem mass spectrometry analysis. RESULTS: We identified a total of 13,414 peptides on 1802 proteins, of which 169 and 47 proteins were significantly differentially expressed at acute and chronic infection phases, respectively. Clustering analysis revealed obvious differences in proteome profiles among all groups. Gene ontology analysis showed that a large number of differentially expressed proteins (DEPs) detected in acute infection were associated with biological binding activity and single-organism processes. KEGG pathway enrichment analysis showed that the majority of these DEPs were involved in disease-related and metabolic pathways. CONCLUSIONS: Our findings revealed global reprogramming of the urine proteome following T. gondii infection, and data obtained in this study will enhance our understanding of the host responses to T. gondii infection and lead to the identification of new diagnostic biomarkers.

Comparative genomic analysis of three geographical isolates from China reveals high genetic stability of Plutella xylostella granulovirus.

In this study, the genomes of three Plutella xylostella granulovirus (PlxyGV) isolates, PlxyGV-W and PlxyGV-Wn from near Wuhan and PlxyGV-B from near Beijing, China were completely sequenced and comparatively analyzed to investigate genetic stability and diversity of PlxyGV. PlxyGV-W, PlxyGV-B and PlxyGV-Wn consist of 100,941bp, 100,972bp and 100,999bp in length with G + C compositions of 40.71-40.73%, respectively, and share nucleotide sequence identities of 99.5-99.8%. The three individual isolates contain 118 putative protein-encoding ORFs in common. PlxyGV-W, PlxyGV-B and PlxyGV-Wn have ten, nineteen and six nonsynonymous intra isolate nucleotide polymorphisms (NPs) in six, fourteen and five ORFs, respectively, including homologs of five DNA replication/late expression factors and two per os infectivity factors. There are seventeen nonsynonymous inter isolate NPs in seven ORFs between PlxyGV-W and PlxyGV-B, seventy three nonsynonymous NPs in forty seven ORFs between PlxyGV-W and PlxyGV-Wn, seventy seven nonsynonymous NPs in forty six ORFs between PlxyGV-B and PlxyGV-Wn. Alignment of the genome sequences of nine PlxyGV isolates sequenced up to date shows that the sequence homogeneity between the genomes are over 99.4%, with the exception of the genome of PlxyGV-SA from South Africa, which shares a sequence identity of 98.6-98.7% with the other ones. No events of gene gain/loss or translocations were observed. These results suggest that PlxyGV genome is fairly stable in nature. In addition, the transcription start sites and polyadenylation sites of thirteen PlxyGV-specific ORFs, conserved in all PlxyGV isolates, were identified by RACE analysis using mRNAs purified from larvae infected by PlxyGV-Wn, proving the PlxyGV-specific ORFs are all genuine genes.

Prevalence and multilocus genotyping of Giardia duodenalis in Tan sheep (Ovis aries) in northwestern China.

Giardia duodenalis is a common intestinal protozoa, which can cause the occurrence of diarrhea, weight loss, and even death in animals or human, this threatens the husbandry industry and public health. It can infect virtually humans and all domestic animals including sheep. Tan sheep is one of the most important sheep breeds, which is short-tailed indigenous sheep breed used for production of high quality meat and pelts in China. However, there are no report regarding the occurrence and multilocus genotyping of G. duodenalis in Tan sheep in northwestern China. Thus, the objective of the present study was to investigate the prevalence and multilocus genotypes of G. duodenalis in Tan sheep. 1014 fecal samples were collected from Tan sheep from Ningxia Hui Autonomous Region, and three loci (ß-giardin (bg), glutamate dehydrogenase (gdh) and triosephosphate isomerase (tpi) genes) were amplified by nested PCR. The prevalence of G. duodenalis in Tan sheep was 14.5% (147/1014), two assemblages (assemblage A, n = 43; and E, n = 90) were detected, including one novel assemblage A at bg locus, one novel assemblage A at tpi locus, and 10 and 11 novel subtypes of assemblage E were detected at the bg and gdh loci, respectively. One MLGs was formed based on sequence variation among the three loci. Moreover, 9 Tan sheep were infected with two assemblages (A and E) based on the three loci. These findings expand the host range of G. duodenalis and revealed genetic diversity of G. duodenalis assemblages in Tan sheep.

Molecular Detection, Multilocus Genotyping, and Population Genetics of Enterocytozoon bieneusi in Pigs in Southeastern China.

Enterocytozoon bieneusi is an important opportunistic pathogen widely distributed in humans and animals that causes diarrhea or fatal diarrhea in immunocompromised hosts. To examine the infection status and molecular characteristics of E. bieneusi in pigs, 725 fecal samples were collected from pigs in six areas of Fujian Province. The E. bieneusi genotypes were identified based on the internal transcribed spacer (ITS) regions of the ribosomal RNA (rRNA) gene by nested PCR, and its population genetics were analyzed by multilocus sequence typing (MLST). The results showed that the infection rate of E. bieneusi was 24.4% (177/725), and 11 known genotypes (EbpC, EbpA, CHN-RR2, KIN-1, CHG7, CHS5, CM11, CHG23, G, PigEBITS, and D) and 2 novel genotypes (FJF and FJS) were identified. All the genotypes were found to be clustered into zoonotic Group 1. Moreover, 52 positive samples were successfully amplified at minisatellite and microsatellite loci and formed 48 distinct multilocus genotypes (MLGs). Further population structure analyses showed strong genetic linkage disequilibrium (LD) and several recombination events (Rm), indicating that E. bieneusi has a clonal population structure. This study is the first to investigate the prevalence and molecular characteristics of E. bieneusi in Fujian Province and could provide baseline data to control E. bieneusi infection in pigs and humans and deepen our understanding of the zoonotic risk of E. bieneusi and its distribution in China.

Prevalence of gastrointestinal parasites in free-range yaks (Bos grunniens) in Gansu Province, Northwest China.

BACKGROUND: Little information about the prevalence of gastrointestinal parasites in yaks (Bos grunniens) in northwest China is available. Therefore, the objective of the study was to quantify faecal egg counts of gastrointestinal parasites (helminths and coccidia) in free-range yaks from Gannan Tibetan Autonomous Prefecture, Gansu Province, Northwest China. RESULTS: Parasites were detected in 290 of 733 (39.56%) faecal samples. The results showed that Strongylidae, Trichuris spp. and Eimeria spp. were detected all year round, Strongyloides papillosus was detected in autumn and summer, and Nematodirus spp. was detected in both autumn and spring. In contrast, Fasciola spp. was only detected in spring. The prevalence rates of parasitic infections in different seasons were significantly different. CONCLUSIONS: To our knowledge, this is the first investigation of gastrointestinal parasites in yaks (Bos grunniens) in Gansu, China. The results demonstrated a high prevalence of gastrointestinal parasitic infections, specifically GN infections, in yaks in GTAP and these infections can cause economic losses to the local cattle industry.

Prevalence and Genetic Identification of Three Entamoeba Species in Pigs in Southeastern China.

Parasitic Entamoeba spp. can infect many classes of vertebrates including humans and pigs. Entamoeba suis and zoonotic Entamoeba polecki have been identified in pigs, and swine are implicated as potential reservoirs for Entamoeba histolytica. However, the prevalence of Entamoeba spp. in pigs in southeastern China has not been reported. In this study, 668 fecal samples collected from 6 different regions in Fujian Province, southeastern China, were analyzed to identify three Entamoeba species by nested PCR and sequencing analysis. The overall prevalence of Entamoeba spp. was 55.4% (370/668; 95% CI 51.6% to 59.2%), and the infection rate of E. polecki ST1 was the highest (302/668; 45.2%, 95% CI 41.4% to 49.0%), followed by E. polecki ST3 (228/668; 34.1%, 95% CI 30.5% to 37.7%) and E. suis (87/668; 13.0%, 95% CI 10.5% to 15.6%). E. histolytica was not detected in any samples. Moreover, the coinfection rate of E. polecki ST1 and ST3 was 25.1% (168/668; 95% CI 21.9% to 28.4%), the coinfection rate of E. polecki ST1 and E. suis was 3.7% (25/668; 95% CI 2.3% to 5.2%), the coinfection rate of E. polecki ST3 and E. suis was 0.3% (2/668), and the coinfection rate of E. polecki ST1, E. polecki ST3, and E. suis was 4.0% (27/668; 95% CI 2.5% to 5.5%). A representative sequence (MK347346) was identical to the sequence of E. suis (DQ286372). Two subtype-specific sequences (MK357717 and MK347347) were almost identical to the sequences of E. polecki ST1 (FR686383) and ST3 (AJ566411), respectively. This is the first study to survey the occurrence and to conduct molecular identification of three Entamoeba species in southeastern China. This is the first report regarding mixed infections with E. suis, E. polecki ST1, and E. polecki ST3 in China. More research studies are needed to better understand the transmission and zoonotic potential of Entamoeba spp.

Immunization With a Live-Attenuated RH:ΔNPT1 Strain of Toxoplasma gondii Induces Strong Protective Immunity Against Toxoplasmosis in Mice.

Toxoplasmosis, one of the most important health-threatening diseases worldwide, is caused by Toxoplasma gondii, which infects a wide range of warm-blooded animals and humans, leading to enormous health and socioeconomic concerns. T. gondii can establish chronic infection to evade the immune response in hosts. Once a chronic infection has been established, the available treatments cannot efficiently control this stage of T. gondii efficiently. Moreover, the available treatments rely only on a few drugs, such as sulfapyridine and pyrimethamine, that tend to have severe side effects. Given these factors, vaccination has been considered to be the most efficient method to prevent and control this disease. However, there is currently lack of effective vaccine available for use to prevent toxoplasmosis apart form Toxovax®, the only available vaccine, which is used in sheep to prevent abortion. To address this problem, we knocked out the NPT1 gene of the type I T. gondii strain using the CRISPR-Cas9 system, constructed a live-attenuated vaccine and evaluated its protective efficacy in a mouse model. Immunization of mice with RH:ΔNPT1 induced a high level of Toxoplasma-specific IgG1, IgG2a and total IgG 42 days after immunization. There was a significant increase in the levels of cytokines in the splenocyte suspensions of RH:ΔNPT1-infected mice, and a mixed Th1/Th2 response was induced in the mice. Remarkably, after heterologous challenges with tachyzoites of the RH, PYS and Pru strains and cysts of the Pru strain by different infection routes, the immunized animals were protected from toxoplasmosis with a 100% survival rate, in both acute and chronic infection. In addition, compared with control mice, the Pru cyst load was clearly reduced in the brains of RH:ΔNPT1-infected immunization-mice. Our study demonstrated that the RH:ΔNPT1 strain was able to evoke strong anti-Toxoplasma immune responses and provide effective protection against parasite strains with different levels of virulence, suggesting that the RH:ΔNPT1 strain may represent a promising live-attenuated vaccine against toxoplasmosis, which is worthy of further evaluation in food-producing animals and in definitive feline host.

Occurrence of Enterocytozoon bieneusi in Chinese Tan sheep in the Ningxia Hui Autonomous Region, China.

Enterocytozoon bieneusi is a zoonotic parasite which is considered to be an opportunistic pathogen of humans and animals. A number of studies have reported E. bieneusi infection in various animals. However, no information is available on the occurrence of E. bieneusi in Tan sheep, a unique indigenous sheep species in the Ningxia Hui Autonomous Region, China. The objectives of the present study were to examine the prevalence and identify the genotypes of E. bieneusi in Tan sheep in China. A total of 1014 fecal specimens of Tan sheep from six farms in the Ningxia Hui Autonomous Region were examined by nested PCR amplification of the internal transcribed spacer (ITS) of nuclear ribosomal DNA. The total prevalence of E. bieneusi was 12.2% (124/1014), ranging from 0.5 to 22.2% on six farms. Sequence analysis identified 10 genotypes of E. bieneusi, including three known genotypes, BEB6, COS-I, and CHG13, and seven novel genotypes designated as NX1 to NX7, which all belonged to group 2 by phylogenetic analysis. This is the first report describing the prevalence of E. bieneusi in Tan sheep, and the new genotypes identified in the current study expand the genotype distribution of E. bieneusi. These findings provide baseline data and have implications for the epidemiology and control of E. bieneusi infection in Tan sheep.

A novel recombinase polymerase amplification (RPA) assay for the rapid isothermal detection of Neospora caninum in aborted bovine fetuses.

The development of a method to rapidly diagnose Neospora caninum infection is highly desirable. Recombinase polymerase amplification (RPA), combined with lateral flow (LF) strips, is a novel approach to rapidly amplify and visualize DNA. We have developed a prototype LF-RPA assay, using primers and a probe that targeted a specific sequence in the N. caninum NC-5 gene. The N. caninum-specific LF-RPA assay was first tested on purified DNA from oocysts and amplified N. caninum DNA to detectable levels in 10 min, at a constant temperature and without the need for an expensive thermocycler. The designed RPA primers and probe displayed 100% specificity for detecting N. caninum without any cross-reaction with DNA from nine related protozoan spp. (eg Toxoplasma gondii, Sarcocystis gigantean, Sarcocystis zuoi, Hammondia hammondi, Hammondia heydorni, Eimeria cylindrica, Plasmodium falciparum, Theileria annulata and Babesia bigemina). Although, LF-RPA assay detected amounts as low as 50 fg of N. caninum DNA, it was nearly 5-fold less sensitive than previously published qPCR and nested PCR assays. We tested the diagnostic performance of the LF-RPA assay for the detection of N. caninum DNA in aborted bovine fetal tissue samples, and compared the results with those obtained from nested PCR. Out of the 75 samples examined, 18 (24%) and 17 (22.6%) tested positive using LF-RPA and nested PCR, respectively. Our results indicate that LF-RPA is a suitable assay for the rapid and reliable detection of N. caninum.

Human impact on the diversity and virulence of the ubiquitous zoonotic parasite Toxoplasma gondii.

A majority of emerging infectious diseases in humans are zoonoses. Understanding factors that influence the emergence and transmission of zoonoses is pivotal for their prevention and control. Toxoplasma gondii is one of the most widespread zoonotic pathogens known today. Whereas only a few genotypes of T. gondii dominate in the Northern Hemisphere, many genotypes coexist in South America. Furthermore, T. gondii strains from South America are more likely to be virulent than those from the Northern Hemisphere. However, it is not clear what factor(s) shaped modern-day genetic diversity and virulence of T. gondii Here, our analysis suggests that the rise and expansion of farming in the past 11,000 years established the domestic cat/mouse transmission cycle for T. gondii, which has undoubtedly played a significant role in the selection of certain linages of T. gondii Our mathematical simulations showed that within the domestic transmission cycle, intermediately mouse-virulent T. gondii genotypes have an adaptive advantage and eventually become dominant due to a balance between lower host mortality and the ability to superinfect mice previously infected with a less virulent T. gondii strain. Our analysis of the global type II lineage of T. gondii suggests its Old World origin but recent expansion in North America, which is likely the consequence of global human migration and trading. These results have significant implications concerning transmission and evolution of zoonotic pathogens in the rapidly expanding anthropized environment demanded by rapid growth of the human population and intensive international trading at present and in the future.

First Report of Seroprevalence and Risk Factors of Neospora caninum Infection in Tibetan Sheep in China.

Neospora caninum is an intracellular protozoan parasite which can cause abortion and stillbirth in ruminants. However, there is no information on Tibetan sheep N. caninum infection in China. A total of 2187 serum samples were collected from Tibetan sheep in the major production areas of Luqu, Maqu, and Tianzhu in Gansu province, and Nyingchi in southeast Tibet, China. All samples were analyzed for the presence of antibodies to N. caninum using a competitive-inhibition enzyme-linked immunoassay. Of the 2187 serum samples, 184 (8.4%, 95% CI 7.3-9.6) were tested N. caninum seropositive. The N. caninum seroprevalence ranged from 4.4% (95% CI 1.4-7.4) to 11.3% (95% CI 8.2-14.4) among different regions, seasons, ages, and pregnancies, and there was no statistical significance among those groups (P > 0.05). Seroprevalence in male (10.8% 69/638) (95% CI 8.4-13.2) was significantly higher than in female (7.4% 115/1549) (OR =1.51, 95% CI 6.1-8.7) (P < 0.01). To our knowledge, this is the first report of N. caninum seroprevalence in Tibetan sheep in China, which provides baseline data for the prevention and control of N. caninum infection in Tibetan sheep.

Characterization of the complete mitochondrial genome of Marshallagia marshalli and phylogenetic implications for the superfamily Trichostrongyloidea.

Marshallagia marshalli (Nematoda: Trichostrongylidae) infection can lead to serious parasitic gastroenteritis in sheep, goat, and wild ruminant, causing significant socioeconomic losses worldwide. Up to now, the study concerning the molecular biology of M. marshalli is limited. Herein, we sequenced the complete mitochondrial (mt) genome of M. marshalli and examined its phylogenetic relationship with selected members of the superfamily Trichostrongyloidea using Bayesian inference (BI) based on concatenated mt amino acid sequence datasets. The complete mt genome sequence of M. marshalli is 13,891 bp, including 12 protein-coding genes, 22 transfer RNA genes, and 2 ribosomal RNA genes. All protein-coding genes are transcribed in the same direction. Phylogenetic analyses based on concatenated amino acid sequences of the 12 protein-coding genes supported the monophylies of the families Haemonchidae, Molineidae, and Dictyocaulidae with strong statistical support, but rejected the monophyly of the family Trichostrongylidae. The determination of the complete mt genome sequence of M. marshalli provides novel genetic markers for studying the systematics, population genetics, and molecular epidemiology of M. marshalli and its congeners.

Comparative proteomic analysis of virulent and avirulent strains of Toxoplasma gondii reveals strain-specific patterns.

Research exploring the proteome of Toxoplasma gondii oocysts has gained momentum over the past few years. However, little is known about the oocyst's protein repertoires that contribute to differential virulence among T. gondii strains. Here, we used isobaric tag for relative and absolute quantitation-based proteomic analysis of oocysts of two T. gondii strains exhibiting the virulent PYS (ToxoDB#9) phenotype versus the less virulent PRU (Type II, ToxoDB#1) phenotype. Our aim was to determine protein expression patterns that contribute to the virulence of a particular phenotype. A total of 2,551 proteins were identified, of which 374 were differentially expressed proteins (DEPs) (|log2 fold change| ≥ 0.58 and P < 0.05). DEPs included 192 increased and 182 decreased proteins. Gene Ontology and KEGG pathway analyses revealed a large number of DEPs enriched in various metabolic processes. Protein interaction network analysis using STRING identified inosine monophosphate dehydrogenase (IMPDH), Bifunctional GMP synthase/glutamine amidotransferase protein, Glucose-6-phosphate 1-dehydrogenase, and Citrate synthase as the top four hubs. Of the 22 virulence proteins commonly expressed in the oocysts of the two strains, 13 and 2 proteins were increased in PYS strain and PRU strain, respectively. Also, 10 and 3 of the 22 identified oocyst wall proteins showed higher expression in oocysts of PRU strain and PYS strain, respectively. These findings revealed new proteomic differences in the oocysts of T. gondii strains of different genotypic backgrounds.

Vaccination with a DNA vaccine encoding Toxoplasma gondii ROP54 induces protective immunity against toxoplasmosis in mice.

Toxoplasma gondii is an obligatory intracellular protozoan, which infects most of the warm-blooded animals, causing serious public health problems and enormous economic losses worldwide. The rhoptry effector protein 54 (ROP54) has been indicated as a virulence factor that promotes Toxoplasma infection by modulating GBP2 loading onto parasite-containing vacuoles, which can modulate some aspects of the host immune response. In order to evaluate the immuno-protective value of ROP54, we constructed a eukaryotic recombinant plasmid expressing T. gondii ROP54 and intramuscularly immunized Kunming mice with this recombinant plasmid against acute and chronic toxoplasmosis. All mice immunized with pVAX-ROP54 elicited a high level of specific antibody responses, a significant increase of lymphocyte proliferation, and a significant level of Th1-type cytokines (IFN-γ, IL-2 and IL-12p70), in addition to an increased production of Th2-type cytokines (IL-4 and IL-10). These results demonstrated that pVAX-ROP54 induced significant cellular and humoral (Th1/Th2) immune responses, which extended the survival time (13.0±1.15days for pVAX-ROP54 vs 6.7±0.48days for pVAX I, 6.8±0.42days for PBS and 6.5±0.53 for blank control) and significantly reduced cyst burden (35.9% for pVAX-ROP54, 1% for pVAX I and 2% for PBS, compared with blank control) of immunized mice. These results indicate that the recombinant ROP54 plasmid can provide partial protection and might be a potential vaccine candidate against acute and chronic toxoplasmosis.