Cross-genera amplification and identification of Colpodella sp. with Cryptosporidium primers in fecal samples of zoo felids from northeast China

Abstract The protozoans include many intracellular human pathogens. Accurate detection of these pathogens is necessary to treat the diseases. In clinical epidemiology, molecular identification of protozoan is considered a more reliable and rapid method for identification than microscopy. Among these protozoans, Cryptosporidium considered being one of the important water-borne zoonotic pathogens and a major cause of a diarrheal disease named cryptosporidiosis in humans, domestic animals, and wild animals. This study was aimed to identify Cryptosporidium in zoo felids (N= 56) belonging to different zoo of China, but accidentlly Colpodella was encountered in the zoo felids sample and phylogenetic data confirmed this unexpected amplification from fecal samples using two-step nested-PCR. Phylogenetic analysis revealed the fact about the specific primers used previously by many researchers and cross-genera amplification. We came to know that genetically sequenced amplicon gives more accurate identification of species. This study suggests more investigation on Colpodella which has been neglected previously but gains the attention of researchers after identified from humans and animals and has been known to correlate with neurological symptoms in patients.

Resumo Os protozoários incluem muitos patógenos humanos intracelulares. A detecção acurada desses patógenos é necessária para tratar as doenças. Na epidemiologia clínica, a identificação molecular de protozoários é considerada o método de identificação mais confiável e rápido do que a microscopia. Entre esses protozoários, o Cryptosporidium é considerado um dos importantes patógenos zoonóticos transmitidos pela água e uma das principais causas de uma doença diarreica denominada criptosporidiose em humanos, animais domésticos e selvagens. Este estudo teve como objetivo identificar Cryptosporidium em zoofelídeos (N = 56) pertencentes a diferentes zoológicos da China, mas acidentalmente Colpodella foi encontrada na amostra de zoofelídeos e os dados filogenéticos confirmaram essa amplificação inesperada de amostras fecais usando nested-PCR em duas etapas. A análise filogenética revelou o fato sobre os primers específicos usados anteriormente por muitos pesquisadores e a amplificação entre gêneros. Ficamos sabendo que o amplicon sequenciado geneticamente fornece uma identificação mais acurada das espécies. Este estudo sugere mais investigação sobre Colpodella, que foi negligenciada anteriormente, mas ganha a atenção dos pesquisadores depois de identificada em humanos e animais e é conhecida por se correlacionar com sintomas neurológicos em pacientes.

Cross-genera amplification and identification of Colpodella sp. with Cryptosporidium primers in fecal samples of zoo felids from northeast China / Amplificação cruzada de gêneros e identificação de Colpodella sp. com iniciadores de Cryptosporidium em amostras fecais de zoofilia do nordeste da China

Abstract The protozoans include many intracellular human pathogens. Accurate detection of these pathogens is necessary to treat the diseases. In clinical epidemiology, molecular identification of protozoan is considered a more reliable and rapid method for identification than microscopy. Among these protozoans, Cryptosporidium considered being one of the important water-borne zoonotic pathogens and a major cause of a diarrheal disease named cryptosporidiosis in humans, domestic animals, and wild animals. This study was aimed to identify Cryptosporidium in zoo felids (N= 56) belonging to different zoo of China, but accidentlly Colpodella was encountered in the zoo felids sample and phylogenetic data confirmed this unexpected amplification from fecal samples using two-step nested-PCR. Phylogenetic analysis revealed the fact about the specific primers used previously by many researchers and cross-genera amplification. We came to know that genetically sequenced amplicon gives more accurate identification of species. This study suggests more investigation on Colpodella which has been neglected previously but gains the attention of researchers after identified from humans and animals and has been known to correlate with neurological symptoms in patients.

Resumo Os protozoários incluem muitos patógenos humanos intracelulares. A detecção acurada desses patógenos é necessária para tratar as doenças. Na epidemiologia clínica, a identificação molecular de protozoários é considerada o método de identificação mais confiável e rápido do que a microscopia. Entre esses protozoários, o Cryptosporidium é considerado um dos importantes patógenos zoonóticos transmitidos pela água e uma das principais causas de uma doença diarreica denominada criptosporidiose em humanos, animais domésticos e selvagens. Este estudo teve como objetivo identificar Cryptosporidium em zoofelídeos (N = 56) pertencentes a diferentes zoológicos da China, mas acidentalmente Colpodella foi encontrada na amostra de zoofelídeos e os dados filogenéticos confirmaram essa amplificação inesperada de amostras fecais usando nested-PCR em duas etapas. A análise filogenética revelou o fato sobre os primers específicos usados ​​anteriormente por muitos pesquisadores e a amplificação entre gêneros. Ficamos sabendo que o amplicon sequenciado geneticamente fornece uma identificação mais acurada das espécies. Este estudo sugere mais investigação sobre Colpodella, que foi negligenciada anteriormente, mas ganha a atenção dos pesquisadores depois de identificada em humanos e animais e é conhecida por se correlacionar com sintomas neurológicos em pacientes.

Cross-genera amplification and identification of Colpodella sp. with Cryptosporidium primers in fecal samples of zoo felids from northeast China.

The protozoans include many intracellular human pathogens. Accurate detection of these pathogens is necessary to treat the diseases. In clinical epidemiology, molecular identification of protozoan is considered a more reliable and rapid method for identification than microscopy. Among these protozoans, Cryptosporidium considered being one of the important water-borne zoonotic pathogens and a major cause of a diarrheal disease named cryptosporidiosis in humans, domestic animals, and wild animals. This study was aimed to identify Cryptosporidium in zoo felids (N= 56) belonging to different zoo of China, but accidentlly Colpodella was encountered in the zoo felids sample and phylogenetic data confirmed this unexpected amplification from fecal samples using two-step nested-PCR. Phylogenetic analysis revealed the fact about the specific primers used previously by many researchers and cross-genera amplification. We came to know that genetically sequenced amplicon gives more accurate identification of species. This study suggests more investigation on Colpodella which has been neglected previously but gains the attention of researchers after identified from humans and animals and has been known to correlate with neurological symptoms in patients.

The effectiveness and safety of thromboprophylaxis in cancer patients based on Khorana score: a meta-analysis and systematic review of randomized controlled trials.

INTRODUCTION: Cancer patients receiving chemotherapy are a high risk of VTE, yet the importance of thromboprophylaxis for cancer patients that are at high risk of developing VTE is still controversial. AIM: To calculate the benefits and harms of thromboprophylaxis, compared to placebo, in ambulatory high-risk cancer patients that are receiving chemotherapy. METHODS: We searched PubMed, Embase, Web of Science, the Cochrane Library, Cochrane Central Register of Controlled Trials, Chinese Biomedical Literature Database, WANFANG Data, Chinese National Knowledge Infrastructure and Chinese Scientific Journal Database for randomized controlled trials (RCTs) describing benefits and harms of thromboprophylaxis. Statistical analysis was performed using Stata software (version 15.1). RESULTS: We included six studies, which contained a total of 3240 cancer patients with thromboprophylaxis and 2874 cancer patients without thromboprophylaxis. Thromboprophylaxis was effective in high-risk patients with two points or higher (RR 0.51, 95% CI 0.36-0.71, I2 = 0.0%, P = 0.526). It was associated with an increase in bleeding events (RR 1.65, 95% CI 1.14-2.40, I2 = 0.0%, P = 0.498) and was mainly efficient in reducing the risk of pulmonary embolism (RR 0.56, 95% CI 0.33-0.96, I2 = 0.0%, P = 0.263). The risk of major (RR 1.85, 95% CI 0.87-3.94, I2 = 0.0%, P = 0.888) and non-major (RR 1.59, 95% CI 0.96-2.62, I2 = 16.3%, P = 0.303) bleeding showed no significant difference with or without thromboprophylaxis. There was no reduction in all-cause mortality with thromboprophylaxis (RR 0.95, 95% CI 0.78-1.18, I2 = 22.0%, P = 0.277). CONCLUSION: Thromoboprophylaxis is effective and safe in cancer patients that are at high risk for developing VTE with chemotherapy.

Retroperitoneal fibrosis secondary to non-urology carcinomas: a clinical and outcome analysis of 97 cases.

PURPOSE: Accumulated evidences have reminded us that since the prognosis of retroperitoneal fibrosis (RPF) secondary to non-urology carcinoma is poor it is essential to distinguish this condition from others. The objective of this study is to investigate the clinical features and outcomes of this infrequent but life-threatening disease. METHODS: A total of 97 patients diagnosed with RPF secondary to non-urology carcinoma were enrolled in this retrospective study. The baseline information, history of carcinoma, clinical presentation, laboratory results, diagnosis, treatment and prognosis of the patients were collected and analyzed. RESULTS: The mean age was 62.32 ± 11.22 years (range 31-85 years). It took 16.72 ± 7.11 days (range 3-34 days) for the patients with originally unexplained hydronephrosis to get a definite diagnosis of RPF secondary to non-urology carcinoma. Imaging-confirmed RPF [confirmed by computed tomography (CT) or magnetic resonance imaging (MRI)] was found in 91 patients. Fifty-seven patients were diagnosed with histological-confirmed malignant RPF. Sixty-six patients received surgical interventions. Cystoscopy and double J tube were performed in 68 cases (102 sides, 172 times) and was failed in 93 times (54.07%). Pyelostomy was performed in 72 patients (92 sides) without a single failure. Furthermore, eighty patients received chemotherapy and/or radiotherapy. The 6-month survival since RPF secondary to non-urology carcinoma was confirmed was 30%. CONCLUSIONS: All patients with newly diagnosed unexplained hydronephrosis should undergo an age-appropriate carcinoma screening. The prognosis of RPF secondary to non-urology carcinoma was dismal. When dealing with hydronephrosis affecting renal function, pyelostomy instead of double J tube catheterization should be performed ahead of time.

Relationship between TLR4 and MCP2 expression levels and habitual abortion.

Habitual abortion is associated with the altered expression of multiple genes. This study was carried out to investigate the relationship between expression of Toll-like receptor 4 (TLR4) and monocyte chemotactic protein 2 (MCP2 or CCL8) and habitual abortion. This was done by detecting and comparing their relative expression in peripheral blood and placental villi of patients and healthy fertile women. Based on our previous research, 85 subjects with habitual abortion (study group) and 40 healthy fertile women (control group), who were admitted to our hospital between June 2013 and December 2014, were enrolled in this study. After these subjects signed written informed consent, peripheral blood samples and villous tissues were collected, from which the total RNA was extracted. The expression of TLR4 and MCP2 was detected with quantitative reverse transcription-polymerase chain reaction, using GAPDH as a reference control. The expression of TLR4 and MCP2 in the peripheral blood and villous tissues of the study group was significantly higher than that of the control group (P < 0.05). A positive correlation was also observed between the changes in expression levels of TLR4 and MCP2. In conclusion, TLR4 and MCP2 expression correlated with the occurrence of habitual abortion. Detecting expression changes in TLR4 and MCP2 in the peripheral blood is a feasible method for predicting the occurrence of abortion in women of child-bearing age.

Serum levels of S-100ß correlate with the clinical status and severity of hypoxic-ischemic encephalopathy in neonates.

The clinical significance of serum S-100ß levels in neonates with hypoxic-ischemic encephalopathy (HIE), as a reference index to assess HIE severity, was evaluated in this study. On the basis of our strict inclusion and exclusion criteria, relevant high-quality case-control studies reporting the association between HIE and S-100ß protein were selected from electronic database searches. The STATA version 12.0 software was used for the statistical analyses. The database search initially retrieved 93 studies (37 in English and 56 in Chinese), and following a multistep screening process, 13 high-quality studies were eventually included in our meta-analysis. The 13 case-control studies included a total of 646 HIE neonates and 381 healthy controls. The results of this meta-analysis revealed that serum S-100ß levels in mild, moderate, and severe HIE neonates were significantly higher than those in healthy controls, and the differences were statistically significant. Importantly, the serum S-100ß levels increased incrementally with HIE severity. Our results support the hypothesis that S-100ß is an important biological indicator of HIE and serum S-100ß levels can be used as a reference index to assess HIE severity.

Expression and clinical significance of PIWIL2 in hilar cholangiocarcinoma tissues and cell lines.

The objective of this study was to explore the relationship between PIWI-like protein 2 (PIWIL2) and clinicopathological charac-teristics and prognosis after radical resection. To accomplish this, we analyzed PIWIL2 expression in hilar cholangiocarcinoma tissues and cell lines. PIWIL2 expression was detected by immunohistochemistry in 41 hilar cholangiocarcinoma samples and 10 control tissues. Western blotting and immunocytofluorescence were used to investigate PIWIL2 expression in the cholangiocarcinoma cell line QBC939 and the bile duct epithelial cell line HIBEpic. Univariate and multivariate surviv-al analyses were performed using the Kaplan-Meier method for hilar cholangiocarcinoma patients who underwent radical resection. PIWIL2 expression was significantly higher in the hilar cholangiocarcinoma tissues and QBC939 cells than in control tissues and HIBEpic cells, respectively (P < 0.05). Poorly and moderately differentiated cholan-giocarcinoma tissues had significantly higher PIWIL2 expression than well-differentiated tissues (P < 0.05). Univariate analysis demonstrated that high PIWIL2 expression was associated with shorter survival time after radical resection (P < 0.05). Multivariate analysis showed that PI-WIL2 expression was an independent prognostic factor after radical re-section of hilar cholangiocarcinoma (P < 0.05). PIWIL2 expression was also associated with tumor-node-metastasis stage and differentiation. PIWIL2 was an independent prognostic factor after radical resection of hilar cholangiocarcinoma.

MEF2A gene mutations and susceptibility to coronary artery disease in the Chinese population.

We investigated mutations and polymorphisms of the coronary artery disease (CAD)-related myocyte enhancer factor 2A (MEF2A) gene in a Chinese population. Polymerase chain reaction-single-strand conformation polymorphism and DNA sequencing were used to detect exon 11 of the MEF2A gene in 210 Hubei patients with CAD and 190 healthy controls. The following mutations were identified: a) a synonymous heterozygous mutation (147191G→T) combined with a 6-base deletion (147123-147128); b) a synonymous heterozygous or homozygous mutation (147191G→T) combined with a 9-base deletion (147123-147131); c) a synonymous mutation (147143G→A); d) a synonymous mutation (147191G→T) combined with an 18-base deletion (147111-147128); and e) a 21-base deletion (147108-147128). Mutations (a) and (b) and a 3-base deletion (147126-147128) with or without the synonymous heterozygous mutation (147191G→T) occurred in more than 1% of controls. However, mutations (c), (d), and (e) were not observed in the control group. The polymorphism in exon 11 of the MEF2A gene was observed in the Chinese population. Six or seven amino acid deletions and synonymous mutations (147143G→A) may be correlated with susceptibility to CAD.

Effects of beraprost sodium on renal function and inflammatory factors of rats with diabetic nephropathy.

Beraprost sodium (BPS) is a prostaglandin analogue. We investigated its effects on rats with diabetic nephropathy. There were 20 rats each in the normal control group (NC), the diabetic nephropathy group (DN), and the BPS treatment group. The rats in DN and BPS groups were given a high-fat diet combined with low-dose streptozotocin intraperitoneal injections. The rats in the BPS group were given daily 0.6 mg/kg intraperitoneal injections of this drug. After 8 weeks, blood glucose, 24-h UAlb, Cr, BUN, hs-CRP, and IL-6 levels increased significantly in the DN group compared with the NC group; however, the body mass was significantly reduced in the DN group compared with the NC group. Blood glucose, urine output, 24-h UAlb, Cr, hs-CRP, and IL-6 levels were significantly lower in the BPS group than in the DN group; the body mass was significantly greater in the DN group. Therefore, we concluded that BPS can improve renal function and protect the kidneys of DN rats by reducing oxidative stress and generation of inflammatory cytokines; it also decreases urinary protein excretion of rats with diabetic nephropathy.

Gene expression profiling of Duchenne muscular dystrophy reveals characteristics along disease progression.

Duchenne muscular dystrophy (DMD) is the most common form of muscular dystrophy with no cure currently available. In this study, using two microarray data sets obtained from the Gene Expression Omnibus database, we conducted a dysfunctional pathway-enrichment analysis and investigated deregulated genes that are specific to different phases of the disease in order to determine pathogenic characteristics in the progression of DMD. We identified 41 and 33 dysfunctional pathways that were enriched with differentially expressed genes in presymptomatic patients and in symptomatic patients, respectively. Over 70% of pathways were shared between both phases and many of them involved the inflammatory process, suggesting that inflammatory cascades were induced soon after the birth of the patients. Further investigation showed that presymptomatic patients performed better with respect to muscle regeneration and cardiac muscle calcium homeostasis maintenance. Neuronal nitric oxide synthase, dihydropyridine receptors, sarcoplasmic/endoplasmic reticulum calcium ATPase, and phospholamban may serve as potential targets for further molecular diagnostic tests. Our results may provide a better understanding for the treatment of DMD.

Leukocyte adhesion--a fundamental process in leukocyte physiology.

Leukocyte adhesion is of pivotal functional importance. The adhesion involves several different adhesion molecules, the most important of which are the leukocyte beta 2-integrins (CD11/CD18), the intercellular adhesion molecules, and the selectins. We and others have extensively studied the specificity and binding sites in the integrins and the intercellular adhesion molecules for their receptors and ligands. The integrins have to become activated to exert their functions but the possible mechanisms of activation remain poorly understood. Importantly, a few novel intercellular adhesion molecules have been recently described, which seem to function only in specific tissues. Furthermore, it is becoming increasingly apparent that changes in integrins and intercellular adhesion molecules are associated with a number of acute and chronic diseases.