[A multicenter study on respiratory pathogen detection with Mycoplasma pneumoniae pneumonia in children].

Objective: To analyze the status of respiratory pathogen detection and the clinical features in children with Mycoplasma pneumoniae pneumonia (MPP). Methods: A prospective, multicenter study was conducted to collect clinical data, including medical history, laboratory examinations and multiplex PCR tests of children diagnosed with MPP from 4 hospitals in China between November 15th and December 20th, 2023. The multiplex PCR results and clinical characteristics of MPP children in different regions were analyzed. The children were divided into severe and mild groups according to the severity of the disease. Patients in the severe group were further divided into Mycoplasma pneumoniae (MP) alone and Multi-pathogen co-detection groups based on whether other pathogens were detected besides MP, to analyze the influence of respiratory pathogen co-detection rate on the severity of the disease. Mann-Whitney rank sum test and Chi-square test were used to compare data between independent groups. Results: A total of 298 children, 136 males and 162 females, were enrolled in this study, including 204 children in the severe group with an onset age of 7.0 (6.0, 8.0) years, and 94 children in the mild group with an onset age of 6.5 (4.0, 7.8) years. The level of C-reactive protein, D-dimer, lactic dehydrogenase (LDH) were significantly higher (10.0 (5.0, 18.0) vs. 5.0 (5.0, 7.5) mg/L, 0.6 (0.4, 1.1) vs. 0.5 (0.3, 0.6) mg/L, 337 (286, 431) vs. 314 (271, 393) U/L, Z=2.02, 2.50, 3.05, all P<0.05), and the length of hospitalization was significantly longer in the severe group compared with those in mild group (6.0 (6.0, 7.0) vs. 5.0 (4.0, 6.0) d, Z=4.37, P<0.05). The time from onset to admission in severe MPP children was significantly shorter than that in mild MPP children (6.0 (5.0, 9.5) vs. 9.0 (7.0, 13.0) d, Z=2.23, P=0.026). All patients completed the multiplex PCR test, with 142 cases (47.7%) MPP children detected with 21 pathogens including adenovirus 25 cases (8.4%), human coronavirus 23 cases (7.7%), rhinovirus 21 cases (7.0%), Streptococcus pneumoniae 21 cases (7.0%), influenza A virus 18 cases (6.0%). The pathogens with the highest detection rates in Tianjin, Shanghai, Wenzhou and Chengdu were Staphylococcus aureus at 10.7% (8/75), adenovirus at 13.0% (10/77), adenovirus at 15.3% (9/59), and both rhinovirus and Haemophilus influenzae at 11.5% (10/87) each. The multi-pathogen co-detection rate in severe MPP children was significantly higher than that in mild MPP group (52.9% (108/204) vs. 36.2% (34/94), χ²=10.62,P=0.005). Among severe MPP children, there are 89 cases in the multi-pathogen co-detection group and 73 cases in the simple MPP group. The levels of LDH, D-dimer and neutrophil counts in the multi-pathogen co-detection group were significantly higher than those in the simple MPP group (348 (284, 422) vs. 307 (270, 358) U/L, 0.8 (0.5, 1.5) vs. 0.6 (0.4, 1.0) mg/L, 4.99 (3.66, 6.89)×109 vs. 4.06 (2.91, 5.65)×109/L, Z=5.17, 4.99, 6.11, all P<0.05). Conclusions: The co-detection rate of respiratory pathogens, LDH and D-dimer in children with severe MPP were higher than those with mild MPP. Among severe MPP children the stress response of children in co-detection group was more serious than that of children with simple MPP.

[Clinical characteristics and vaccination status of SARS-CoV-2 Omicron variant infected children].

Objective: To investigate the clinical characteristics and vaccination status of SARS-CoV-2 Omicron variant infected children. Methods: A total of 105 children infected with Omicron variant admitted to Tianjin Haihe Hospital (designated referral hospital for SARS-CoV-2 infection in Tianjin) from January 8, 2022 to February 3 were included for a retrospective study. The cases were divided into pneumonia group and non-pneumonia group according to chest imaging. Based on the doses of inactivated SARS-CoV-2 vaccine, the children who completed SARS-CoV-2 antibody test within 3 days after hospitalization were divided into 2 dose group and<2 dose group.Rank sum test and Chi-square test were used for the comparison between the groups. Results: The age of these 105 children was 10 (8, 11) years on admission, 53 children were males and 52 were females. Eighty-seven cases (82.9%) had mild symptoms, 13 cases (12.4%) had common symptoms and 5 cases (4.8%) were asymptomatic. Ninety-one cases (86.7%) completed 2 doses vaccination. The clinical symptoms were characterized by cough (74 cases, 70.5%), fever (58 cases, 55.2%), sore or dry throat (34 cases, 32.4%), nasal congestion (28 cases, 26.7%), rhinorrhea (23 cases, 21.9%). None of the children received antivirals, steroids, immunosuppressant or oxygen therapy. Seventy-six cases(72.4%) received traditional Chinese medicine treatment. The pneumonia group had a higher rate of positive SARS-CoV-2 IgG within 1 day after admission (13/13 vs. 87.0% (80/92), χ2=42.81, P<0.001) than the non-pneumonia group. Among the 62 children who had serial SARS-CoV-2 antibody tests within 3 days after hospitalization, Compared to the<2 dose group, the 2 dose group had a higher rate of nucleic acid conversion within 16 days after onset and a higher rate of positive SARS-CoV-2 IgG 1 day after admission and 3 days after hospitalization (96.4% (54/56) vs. 4/6, 100.0% (56/56) vs. 2/6, 100.0% (56/56) vs. 3/6, all P<0.05). Conclusions: Most children infected with Omicron variant have mild symptoms, mainly respiratory infection symptoms. The proportion of SARS-CoV-2 antibody IgG positive in children who have received 2 doses of inactivated SARS-CoV-2 vaccines is higher,and the time of whose nucleic acid conversion may be shortened.

[A multicenter retrospective study on the etiology of necrotizing pneumonia in children].

Objective: To investigate the etiology of necrotizing pneumonia (NP) in children and the clinical characteristics of NP caused by different pathogens in China. Methods: A retrospective, case-control study was performed in children with NP who were admitted to 13 hospitals in China from January 2008 to December 2019. The demographic and clinical information, laboratory data, etiological and radiological findings were analyzed. The data were divided into three groups based on the following years: 2008-2011, 2012-2015 and 2016-2019, and the distribution characteristics of the pathogens in different period were compared. Meanwhile, the pathogens of pediatric NP in the southern and northern China were compared. And the clinical characteristics of the Mycoplasma pneumoniae (MP) NP and the bacterial NP were also compared. T-test or Mann-Whitney nonparametric test was used for comparison of numerical variables, and χ2 test was used for categorical variables. Results: A total of 494 children with NP were enrolled, the median ages were 4.7 (0.1-15.3) years, including 272 boys and 222 girls. Among these patients, pathogens were identified in 347 cases and the pathogen was unclear in the remaining 147 cases. The main pathogens were MP (238 cases), Streptococcus pneumoniae (SP) (61 cases), Staphylococcus aureus (SA) (51 cases), Pseudomonas aeruginosa (13 cases), Haemophilus influenzae (10 cases), adenovirus (10 cases), and influenza virus A (7 cases), respectively. MP was the most common pathogen in all three periods and the proportion increased yearly. The proportion of MP in 2016-2019 was significantly higher than that in 2012-2015 (52.1% (197/378) vs. 36.8% (32/87), χ2=6.654, P=0.010), while there was no significant difference in the proportion of MP in 2012-2015 and that in 2008-2011 (36.8% (32/87) vs. 31.0% (9/29), χ²=0.314, P=0.575).Regarding the regional distribution, 342 cases were in the southern China and 152 in the northern China. Also, MP was the most common pathogen in both regions, but the proportion of MP was higher and the proportion of SP was lower in the north than those in the south (60.5% (92/152) vs. 42.7% (146/342), χ2=13.409, P<0.010; 7.9% (12/152) vs. 14.3% (49/342), χ2=4.023, P=0.045). Comparing the clinical characteristics of different pathogens, we found that fever and cough were the common symptoms in both single MP and single bacterial groups, but chest pain was more common (17.0% (34/200) vs. 6.1% (6/98), χ2=6.697, P=0.010) while shortness of breath and wheezing were less common in MP group (16.0% (32/200) vs. 60.2% (59/98), χ2=60.688, P<0.01; 4.5% (9/200) vs. 21.4% (21/98), χ2=20.819, P<0.01, respectively). The white blood cell count, C-reactive protein and procalcitonin in the bacterial group were significantly higher than those in the MP group (14.7 (1.0-67.1)×109/L vs. 10.5 (2.5-32.2)×109/L, 122.5 (0.5-277.3) mg/L vs. 51.4 (0.5-200.0) g/L, 2.13 (0.05-100.00) µg/L vs. 0.24 (0.01-18.85) µg/L, Z=-3.719, -5.901 and -7.765, all P<0.01). Conclusions: The prevalence of pediatric NP in China shows an increasing trend during the past years. MP, SP and SA are the main pathogens of NP, and the most common clinical symptoms are fever and cough. The WBC count, C-reactive protein and procalcitonin in bacterial NP are significantly higher than those caused by MP.

[Consistency of peripheral whole blood and venous serum procalcitonin in children: a multicenter parallel controlled study].

Objective: To explore the consistency of peripheral whole blood and venous serum procalcitonin (PCT) levels, and the value of peripheral whole blood PCT in evaluating pediatric bacterial infection. Methods: This multicenter cross-sectional parallel control study was conducted in 11 children's hospital. All the 1 898 patients older than 28 days admitted to these hospitals from March 2018 to February 2019 had their peripheral whole blood and venous serum PCT detected simultaneously with unified equipment, reagent and method. According to the venous serum PCT level, the patients were stratified to subgroups. Analysis of variance and chi-square test were used to compare the demographic characteristics among groups. And the correlation between the peripheral blood and venous serum PCT level was investigated by quantitative Pearson correlation analysis.The PCT resultes were also converted into ranked data to further test the consistency between the two sampling methods by Spearman's rank correlation test. Furthermore, the ranked data were converted into binary data to evaluate the consistency and investigate the best cut-off of peripheral blood PCT level in predicting bacterial infection. Results: A total of 1 898 valid samples were included (1 098 males, 800 females),age 27.4(12.2,56.7) months. There was a good correlation between PCT values of peripheral whole blood and venous serum (r=0.97, P<0.01). The linear regression equation was PCTvenous serum=0.135+0.929×PCTperipheral whole blood. However, when stratified to 5 levels, PCT results showed diverse and unsatisfied consistency between the two sampling methods (r=0.51-0.92, all P<0.01). But after PCT was converted to ordinal categorical variables, the stratified analysis showed that the coincidence rate of the measured values by the two sampling methods in each boundary area was 84.9%-97.1%. The dichotomous variables also showed a good consistency (coincidence rate 96.8%-99.3%, Youden index 0.82-0.89). According to the severity of disease, the serum PCT value was classified into 4 intervals（<0.5、0.5-<2.0、2.0-<10.0、≥10.0 µg/L）, and the peripheral blood PCT value also showed a good predictive value (AUC value was 0.991 2-0.997 9). The optimal cut points of peripheral whole blood PCT value 0.5、1.0、2.0、10.0 µg/L corresponding to venous serum PCT values were 0.395, 0.595, 1.175 and 3.545 µg/L, respectively. Conclusions: There is a good correlation between peripheral whole blood PCT value and the venous serum PCT value, which means that the peripheral whole blood PCT could facilitate the identification of infection and clinical severity. Besides, the sampling of peripheral whole blood is simple and easy to repeat.

[Modified posteromedial approach via lateral side of flexor hallucis longus for the treatment of posterior Pilon fracture].

Objective: To compare the clinical efficacy and the level of muscle and soft tissue damage between modified posteromedial approach via lateral side of flexor hallucis longus and modified posteromedial approach in the treatment of posterior Pilon fracture. Methods: Total of 43 patients (27 males and 16 females, aged from 19 to 71 years) diagnosed with posterior Pilon fracture from June 2016 to June 2018 in Foshan Hospital of Traditional Chinese Medicine were randomly divided into observation group (modified posteromedial approach via lateral side of flexor hallucis longus, 21 cases) and control group (modified posteromedial approach, 22 cases) according to the operation approach. The preoperative waiting time, intraoperative time, intraoperative blood loss, hospitalization time and the complications were recorded and compared between the two groups. The differences of blood creatine kinase (CK), myoglobin (Myo) and C-reactive protein (CRP) at different time points before and after operation were compared between the two groups to elevate the level of muscle and soft tissue damage. The fracture reduction qualities of the two groups were compared by Burwell-Charnley criteria. The differences of fracture healing time, range of motion of metatarsophalangeal joint of the great toe (MTP-ROM), ankle range of motion (Ankle-ROM), American Orthopaedic Foot & Ankle Society (AOFAS) score and visual analogue scale (VAS) score of pain were compared between the two groups at the last follow-up. Results: The observation group and the control group were followed-up for (19±6) months and (16±8) months, respectively; there was no significant difference between the two groups (P>0.05). There were no significant differences in preoperative waiting time, intraoperative blood loss, hospitalization time and fracture healing time between the two groups (all P>0.05). At the last follow-up, there was no significant difference in the MTP-ROM and Ankle-ROM between the two groups (both P>0.05); the AOFAS score of the observation group was 88.2±7.8 and it was 84.5±7.6 in the control group (P>0.05); the VAS score of the observation group was (0.9±1.0) and it was (1.3±0.8) in the control group(P>0.05). Anatomical reduction rate in observation group was higher than that in control group (90.5% vs 81.8%, P>0.05). The operation time in the observation group was (87±16) min and it was (98±11) min in the control group (P<0.05). CK, Myo and CRP were increased in both groups after surgery, but there was no statistical significance between groups at the same time point (all P>0.05). There was no nerve injury in the observation group, while 2 cases (9.0%) of nerve paralysis occurred in the control group. No incision infection and checkrein deformity of the Hallux was found in the two groups. Conclusion: The modified posteromedial approach via lateral side of flexor hallucis longus can obtain good operative field exposure, and does not increase muscle and soft tissue injury, with shorter operative time and fewer complications, without nerve injury and checkrein deformity, it is a safe approach for the treatment of posterior Pilon fracture.

[Clinical analysis of 96 cases of congenital cystic lung lesions in children].

Objective: To summarize the diagnosis, clinical manifestations, treatment and prognosis of congenital cystic lung lesions. Methods: A retrospective study described the clinical course of 96 patients (46 female and 50 male) diagnosed with congenital cystic lung lesions treated at the Tianjin Children's Hospital from January 2010 to March 2019. The clinical findings, imaging examinations, pathological findings, treatment and follow-up were analyzed. Results: Totally 96 patients (aged from 4 days to 13 years) with congenital cystic lung lesions were included in this study. Eighty-six patients (90%) were diagnosed when they had cough and fever symptoms. Forty (42%) patients exhibited congenital cystic adenomatoid malformation, 30 underwent surgical excision, two were at emergency operations and one dead. There were 12 (13%) patients with pulmonary sequestration and four were surgical treated. Twelve (13%) patients with bronchogenic cyst were included and 4 were surgically treated. There were 3 (3%) patients with congenital lobar emphysema and one was surgically treated. Another patient with pneumothorax was operated in other hospital 2 months after discharge. Twenty-nine (30%) patients with unclassified congenital cystic lung lesions could not be definitively diagnosed by CT. Some of them were difficult to be distinguished from necrotizing pneumonia. Finally, 2 patients were diagnosed as necrotizing pneumonia after 6, 10 months follow-up. After operation 37 out of 39 patients recovered well. Conclusions: The diagnosis of congenital pulmonary cystic disease depend on imaging and pathological examination. Most patients are diagnosed when they have respiratory tract infection. The main clinical manifestations are cough and fever. The prognosis of operative management is good.

[Prognostic significance of CLL-IPI for Chinese patients with chronic lymphocytic leukemia].

Objective: To validate the prognostic value of chronic lymphocytic leukemia-international prognostic index (CLL-IPI) for Chinese CLL patients. Methods: Two hundred and fifteen CLL patients who were initially diagnosed and treated in Jiangsu Province Hospital from January 2002 to November 2017 were included in the retrospective analysis. Risk stratification and prognosis were evaluated by CLL-IPI scoring system. Results: ①Of the 215 patients, 143 were males and 72 were females, with a median age of 60 (16-85) years old. The median treatment-free survival (TFS) and overall survival (OS) was 16 months (4-24 months) and 180 months (145-215 months), respectively. ② The median TFS for low (n=60), intermediate (n=50), high (n=45) and very high risk group (n=60) according to the CLL-IPI scoring system was 56, 15, 12 and 5 months, respectively (P<0.001). ③ The median follow-up was 48 months (1-192 months). The median OS for low risk group was not reached and for intermediate, high, and very high risk group was 180, 89 and 74 months, respectively. The estimated 5-year OS rate was 97.6%, 83.7%, 67.8% and 55.2%, respectively (P<0.001). ④ Multivariate analysis indicated that unmutated immunoglobulin heavy chain variable region (IGHV) gene and ß(2)-microglobulin>3.5 mg/L(P<0.001) were independent prognostic factors of TFS, while TP53 deletion and/or mutation(P=0.008), unmutated IGHV (P=0.017) and age>65 years(P=0.045) were independent prognostic factors of OS. Conclusion: CLL-IPI is the powerful tool for risk stratification in Chinese CLL patients.

Short communication: Comparative proteomic analysis of the lactating and nonlactating bovine mammary gland.

The objective of this study was to determine how bovine mammary protein profiles vary during lactation and the dry period. Three lactating and 3 nonlactating cows were selected for mammary gland tissue sampling. Compared with the mammary proteins in nonlactating cows, a total of 60 differentially expressed proteins (DEP, including 57 upregulated and 3 downregulated) were identified in lactating cows using 2-dimensional difference gel electrophoresis combined with mass spectrometry. These DEP included enzymes and proteins associated with various macromolecular metabolic processes, and appeared to promote the increased metabolic activity associated with milk synthesis and secretion. The increased DEP were primarily related to initiation, maintenance, and involution of lactation, and included proteins involved in glycolysis/gluconeogenesis, the tricarboxylic acid cycle, the pentose phosphate pathway, oxidative phosphorylation, aminoacyl-transfer RNA biosynthesis, and fatty acid biosynthesis. Identified DEP were further validated by real-time, reverse-transcription PCR and Western blot. Five new DEP associated with lactation were uniquely identified. This work provided some protein-associated insights to facilitate further investigation of the mechanisms underlying lactation in dairy cows.

Anti-Aging Effect of Riboflavin Via Endogenous Antioxidant in Fruit fly Drosophila Melanogaster.

OBJECTIVES: This study investigated the effect of riboflavin on aging in Drosophila melanogaster (fruit fly). DESIGN: Experimental study. SETTING: Naval Medical Research Institute. PARTICIPANTS: Fruit fly Drosophila melanogaster. INTERVENTION: After lifelong supplement of riboflavin, the lifespan and the reproduction of fruit flies were observed. Hydrogen peroxide (H2O2) was used to mimic oxidative stress damage to fruit flies and the survival time was recorded. MEASUREMENTS: The activity of copper-zinc-containing superoxide dismutase (SOD1), manganese containing SOD (SOD2) and catalase (CAT) and lipofuscin (LF) content were determined. RESULTS: Riboflavin significantly prolonged the lifespan (Log rank χ2=16.677, P<0.001) and increased the reproductive capacity (P<0.01 for day 15; P<0.05 for day 30) of fruit flies by lifelong supplement. The survival time of fruit flies damaged by H2O2 was significantly prolonged (Log rank χ2=15.886, P<0.001), the activity of SOD1 (P<0.01) and CAT (P<0.01) was enhanced, and the accumulation of LF (P<0.01) was inhibited by riboflavin supplement. CONCLUSION: Riboflavin prolonged the lifespan and increased the reproduction of fruit flies through anti-oxidative stress pathway involving enhancing the activity of SOD1 and CAT and inhibiting LF accumulation. Riboflavin deserves more attention for slowing human aging.

[Retrospective analysis of operative methods and complications in 29 patients with carotid body tumor].

Objective:To analyze the operative methods and complications for surgical resection of carotid body tumor (CBT). Method:Clinical data of 29 patients underwent CBT resection from Jan 2007 to Apr 2015 were retrospectively studied. Result:Five out of 29 patients got bilateral CBT, the others had unilateral lesions, totally 34 CBT resection were performed. Tumor completely dissected without carotid artery clamping and reconstruction in 18 procedures, tumor resection combined with external carotid artery resection in 16 procedures, 9 out of these 16 procedures combined with internal carotid artery resection. The internal carotid artery was reconstructed with autologous greater saphenous vein in 4 procedures, with artificial graft in 3 procedures, and internal carotid artery ligation without reconstruction in 2 procedures. There was no patient death during perioperative period, cerebral infarction happened in 1 patient and cranial nerve injury occurred in 12 cases. Conclusion:Carotid artery resection and reconstruction is very common during CBT resection, surgeon must be familiar with the methods of carotid artery reconstruction, and the most common complication of carotid body tumor resection is the cranial nerve injury.

Establishment and characterization of a fish-cell line from the brain of Japanese flounder Paralichthys olivaceus.

A new brain-cell line derived from Japanese flounder Paralichthys olivaceus (POBC) was established. POBC was subcultured for 67 passages over the course of 420 days. The cultured cells were primarily epithelioid-like. Chromosome analysis revealed the cell line to possess the normal P. olivaceus diploid karyotype of 2n = 48t (telocentric chromosomes). The cells exhibited the astrocyte marker glial fibrillary acidic protein by immunocytochemistry, and significant fluorescent signals were observed when the cells were transfected with green fluorescent protein reporter plasmid. The established POBC would be ideal material for the study of function of fish ependyma, the central neuroendocrine system and endocrine disruptors in the marine environment.

Clinical efficacy of montelukast sodium in treating infantile wheezing.

OBJECTIVES: The efficacy and safety of a single-dose of Montelukast sodium for treating virus-related infantile wheezing are investigated in this study. PATIENTS AND METHODS: A prospective, open, randomized, controlled study was carried out on 595 cases of infants who exhibited wheezing after a respiratory syncytial virus infection. Treatment with Montelukast sodium was provided over the course of 12 weeks. The clinical efficacy of Montelukast sodium was determined based on the clinical symptom score, tidal breathing lung function, and short-acting bronchodilator usage, as well as infantile asthma diagnosis rate change at the 4th and 12th week after the administration of the treatment. The adverse reactions were also observed, and a control group was set. The mean age of the 595 patients with infantile wheezing was 10.82 months ± 4.22 months. Among these patients, 45.9% (273 out of 595) had a family history of asthma, 30.6% (182 out of 595) had allergic rhinitis, 23.9% (142 out of 595) increased peripheral blood eosinophilia, 6.1% (36 out of 595) exhibited total IgE increase, 40.0% (238 out of 595) had a recurrent history of wheezing, and 64.0% (381 out of 595) had a family history of eczema. RESULTS: After 12 weeks of treatment, the clinical symptom scores significantly improved. Significant differences in the cough, wheezing, and motility scores were observed before and after the treatment (p < 0.05). TPTEF/TE and VPEF/VE significantly improved (p < 0.05) after the treatment. The asthma diagnosis rate was 9.6% (57 out of 595). At four weeks after treatment, various indicators correspondingly improved. Twenty-nine (4.9%) patients exhibited adverse reactions, 55.2% exhibited excitation, 20.7% suffered from insomnia, 10.3% had headaches, 3.4% had erythra, 3.4% suffered from abdominal pain, and 3.4% exhibited an increased glutamic-pyruvate transaminase level. The symptoms of eczema were relieved to some extent, and the symptoms of rhinitis became less serious. Significant differences were observed in the number of wheezing attacks, annual number of days hospitalized, annual number of days when ß2AG was utilized, and lung function improvement (p < 0.05). CONCLUSIONS: Montelukast sodium is clinically effective in treating virus-related wheezing, and clinical application for 4 weeks to 12 weeks can effectively relieve the symptoms of wheezing, improve lung function, and reduce the incidence rate of infantile asthma. Montelukast sodium also causes few adverse reactions.

Screening of genes expressed in vivo after infection by Vibrio anguillarum M3.

AIMS: Genes uniquely expressed in vivo may contribute to the overall pathogenicity of an organism and are likely to serve as potential targets for the development of new vaccine. This study aims to screen the genes expressed in vivo after Vibrio anguillarum infection by in vivo-induced antigen technology (IVIAT). METHODS AND RESULTS: The convalescent-phase sera were obtained from turbot (Scophthalmus maximus) survived after infection by the virulent V. anguillarum M3. The pooled sera were thoroughly adsorbed with M3 cells and Escherichia coli BL21 (DE3) cells. A genomic expression library of M3 was constructed and screened for the identification of immunogenic proteins by colony immunoblot analysis with the adsorbed sera. After three rounds of screening, 19 putative in vivo-induced (ivi) genes were obtained. These ivi genes were catalogued into four functional groups: regulator/signalling, metabolism, biological process and hypothetical proteins. Three ivi genes were insertion-mutated, and the growth and 50% lethal dose (LD(50) ) of these mutants were evaluated. CONCLUSIONS: The identification of ivi genes in V. anguillarum M3 sheds light on understanding the bacterial pathogenesis and provides novel targets for the development of new vaccines and diagnostic reagents. SIGNIFICANCE AND IMPACT OF THE STUDY: To the best of our knowledge, this is the first report describing in vivo-expressed genes of V. anguillarum using IVIAT. The screened ivi genes in this study could be new virulent factors and targets for the development of vaccine, which may have implications for the development of diagnostic regents.

Acute toxicity of a single dose DATR, recombinant soluble human TRAIL mutant, in rodents and crab-eating macaques.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been reported to possess activity of inducing apoptosis in variety of tumor cells in preclinical models. Several mutational versions of TRAIL have been studied as promising agents for cancer therapy and the recombinant soluble human TRAIL mutant (DATR) is one of them. The objective of the present study was to provide possible toxic target organs and proposal non-toxic dose level of DATR for clinical usage. Rodents and crab-eating macaques were used to estimate potential adverse effects of DATR following a single dose administration. The median lethal dose (LD(50)) of intravenous injection to rats and mice was determined as 262.0 and 1018.0 mg/kg b.w., respectively. The LD(50) of intraperitoneal administration to mice was found to be 1432.1 mg/kg b.w. The main changes in macaques were found in the following aspects. Hematology analysis revealed an obvious decrease of red blood cell count (RBC), hemoglobin (HB) and hematocrit (HCT) after injection. Serum biochemical analysis showed an apparent increase of alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), blood urea nitrogen (BUN) and creatinine (Crea). Furthermore, inflammatory cell infiltrate in liver and kidney was found by microscope. All the disorders suggested that liver, renal and hematological systems might be the target effectors of toxic effect induced by DATR. Based on the results of this study, the no observed-adverse-effect level (NOAEL) and the lowest observed-adverse-effect level of DATR in macaques are 90.0 and 135.0 mg/kg b.w., respectively.