OBJECTIVES: To detect prfA and hly toxin genes of Listeria monocytogenes using polymerase chain reaction (PCR) and colloidal gold technology. METHODS: L. monocytogenes DNA was extracted by boiling method. With prfA and hly of L. monocytogenes as the target genes, the 5' ends of upstream and downstream primers of prfA gene were labeled with 6-FAM and biotin, and the 5' ends of upstream and downstream primers of hly gene were labeled with digoxin and biotin, respectively, to establish the toxin gene detection method. Using cloning transformation, sequencing analysis, cloning of positive control products, the detection kid was developed and its specificity, sensitivity, reproducibility and stability were tested, followed by verification with sample testing. RESULTS: The concentration of L. monocytogenes DNA extracted by boiling method was 148.81±0.97 ng/μL, and the A₂₆₀/A₂₈₀ ratio ranged from 1.8 to 2.0. The PCR products showed a 100% homology with the gene sequences in GenBank database after cloning, transformation and sequencing. The colloidal gold strip yielded positive results only for L. monocytogenes samples without cross-reactions with Staphylococcus aureus, Escherichia coli or Bacillus cereus, and its minimum detection limit was 10^-2 ng/μL, demonstrating a 10-fold greater sensitivity of the test than agarose gel electrophoresis. The test also showed good reproducibility of the results when performed by different operators with good stability of the test strips after storage for 6 to 12 months. The test results showed that this kit could accurately and quickly detect L.monocytogenes in the test samples. CONCLUSIONS The detection kit developed in this study can simultaneously detect prfA and hly toxin genes of L. monocytogenes with good specificity, sensitivity, reproducibility and stability for use in food safety inspection.
Listeria monocytogenes/isolation & purification* ; Gold Colloid ; Bacterial Toxins/genetics* ; Polymerase Chain Reaction/methods* ; Hemolysin Proteins/genetics* ; Bacterial Proteins/genetics* ; DNA, Bacterial/genetics* ; Food Microbiology ; Heat-Shock Proteins

Objective: To study the efficacy and safety of sofosbuvir and daclatasvir regimens for patients who received kidney transplantation (KT) with hepatitis C virus (HCV) infection. Methods: This study enrolled a prospective cohort of consecutive KT patients with HCV infection from March 2016 to January 2018 in the hepatology Department of the Second Hospital of Shandong University. They were given sofosbuvir combined with daclatasvir, with or without ribavirin. The course of treatment was 12 weeks or 24 weeks. Clinical assessment, conventional liver and kidney biochemical parameters, hemoglobin, serum HCV RNA, as well as the types of immunosuppressive drugs and their doses were assessed routinely as follows: at the beginning of treatment; 2, 4, and 8 wk post treatment; at the end of treatment (EOT); and at 12, 24 wk after the therapy was completed. Adverse events and adjustment of anti-rejection drugs were surveilled during the treatment period. Results: A total of 13 patients were enrolled. All patients were naive to treatment. Their mean age was 46.84±7.79 years. There were 10 males and 3 females, 3 patients had cirrhosis (1 cases had decompensated cirrhosis), 10 patients had no cirrhosis. They were infected with HCV genotype 1 (6/13 GT1b), genotype 3 (2/13 GT3a) and genotype 6 (3/13 GT6a), and genotype 2 (2/13 GT2a). Twelve patients′ estimated glomerular filtration rate (eGFR) was > 30 ml/min per 1.73 m² at the beginning of treatment, 1 patient′s eGFR was <30 ml/min·1.73 m^2; 9 patients received 12 wk therapy, 4 patients received 24 wk therapy. Twelve patients had undetectable viral load by week 4 of treatment. All patients had undetectable HCV viral load at the end of treatment. Sustained virological response (SVR) 12 rate was achieved in 100% (13/13) of the recipients. The basic renal function remained stable during the course of treatment. No serious adverse events were observed during the treatment. Antiviral therapy was not discontinued due to side effects in any patient. Conclusions Sofosbuvir and daclatasvir for treatment of KT patients with HCV infection are highly effective and safe.

Objective To study the alterations of renal function in patients with acute ischemic stroke and diabetes and to analyze the risk factors of contrast-induced nephropathy (CIN) after digital subtraction angiography (DSA).Methods Eight hundred and seventy-one cerebral infarction patients with diabetes who underwent DSA were selected in the Third Affiliated Hospital of Chongqing Medical University and Nanjing Brain Hospital Affiliated to Nanjing Medical University from August 2012 to August 2016.The patients were divided into diabetic group (n =178) and non-diabetic group (n =693).The alterations of renal function and the incidence rate of CIN were observed between two groups 3 days after DSA.Univariate analysis and multi-factor Logistic regression analysis were used to analyze the independent risk factors of CIN.Results The levels of estimated glomerular filtration rate (eGFR,ml · min-1 · 1.73 m-2) in diabetic group at 1,2 and 3 days after DSA(81.94 ±9.38,75.36 ±8.21,84.43 ±9.72) were lower than that in non-diabetic group (84.62 ± 10.06,79.08 ±9.84,87.62 ± 10.15,t =3.213,4.645,3.772,all P ＜ 0.05).The levels of serum creatinine (Scr) and cystatin (CysC) in diabetic group at 1,2 and 3 days after DSA (Scr:85.63 ±9.83,92.37 ±10.07,83.43 ±9.07;CysC:1.08 ±0.12,1.35 ±0.14,0.95 ±0.10) were higher than that in non-diabetic group (Scr:81.36 ± 8.98,87.84 ± 9.85,80.31 ± 8.64,t =5.548,5.448,4.253;CysC:0.97 ±0.11,1.21 ±0.12,0.88 ±0.09;t =11.677,13.400,9.043;all P ＜ 0.05).The incidence rate of CIN in diabetic group (25.84％ (46/178)) was higher than that in nondiabetic group (7.07％ (49/693),x2 =51.358,P =0.001).Multi-factor Logistic regression analysis showed allergies,plasma brain natriuretic peptide,heart failure,the original renal insufficiency,NIHSS score,contrast agent dosage,preoperative eGFR,preoperative Scr and preoperative CysC were the independent risk factors of CIN in cerebral infarction patients with diabetes.Conclusions The renal function decreased significantly and the incidence rate of CIN was high in cerebral infarction patients with diabetes after DSA.Allergies,plasma brain natriuretic peptide,heart failure,the original renal insufficiency,contrast agent dosage,preoperative eGFR,preoperative Scr and preoperative CysC were the independent risk factors of CIN in cerebral infarction patients with diabetes.

Objective To investigate any potential and independent demographic and serologic risk factors contributing to bone destruction in patients with rheumatoid arthritis ( RA) . Methods A total of 445 patients with RA were recruited in this study. Three autoantibodies including rheumatoid factor ( RF) , anti-cyclic citrullinated peptide antibody ( anti-CCP antibody) and anti-citrullinated alpha-enolase peptide 1 antibody ( anti-CEP-1 antibody) were quantified by using specific ELISA kits. The hand radiographs of all subjects were graded by using the modified Sharp/van der Heijde score ( Sharp score) . The potential and in-dependent risk factors were assessed by using univariate linear regression analyses and the stepwise multiple regression analysis, respectively. Results Based upon the univariate regression analyses, 7 covariates were identified as the potential risk factors for bone destruction in patients with RA, which were female (β=0. 100, P=0. 035), longer disease duration (β=0. 498, P=3. 26×10-29), RF (β=0. 096, P=0. 042), younger age at onset (β=-0. 312, P=1. 60 × 10-11 ), anti-CCP antibody positive (β=0. 202, P=1.74×10-5), anti-CEP-1 antibody positive (β=0.148, P=0.017) and positive for either anti-CCP or anti-CEP-1 antibodies (β=0. 157, P=1. 42×10-3). However, smoking (β=-0. 121, P=0. 018) were identi-fied as the potential protective factors. The multiple regression analysis indicated that the longer disease du-ration (P=2. 24×10-15) and anti-CCP antibody positive (P=0. 012) were independent risk factors for bone destruction. Conclusion Female, longer disease duration, younger age at onset, RF, anti-CCP and anti-CEP-1antibodies are potential risk factors for bone damage in patients with RA. Moreover, longer disease du-ration and anti-CCP antibody are two independent risk factors contributing to bone destruction in RA.

OBJECTIVE:To observe the clinical efficacy and safety of Bromfenac sodium eyed drops in the treatment of xe-rophthalmia. METHODS:80 patients with xerophthalmia were randomly divided into observation group and control group,with 40 cases in each group. Control group was given 0.1% Sodium hyaluronate,one drop,qd;observation group was given 0.1% Sodi-um hyaluronate eye drops,one drop,bid. Both groups received 14 d of treatment. The subjective symptom and sign,the time of lacrimal film break-up,fluorescent staining score and schemer test were observed in 2 groups before treatment,3,7,14 d after treatment. Clinical efficacy and the occurrence of ADR were observed in 2 groups. RESULTS:There was no statistical significance in subjective symptom and sign,the time of lacrimal film break-up,fluorescent staining score and tear between 2 groups before treatment,3,7 d after treatment(P>0.05). 14 d after treatment,subjective symptom and sign and fluorescent staining score of 2 groups were decreased significantly,and the time of lacrimal film break-up was prolonged and secretion was increased significant-ly;the observation group was significantly better than the control group,with statistical significance (P<0.05). The cure rate of observation group was 60.0%,which was significantly higher than that of control group(35.0%),with statistical significance(P<0.05). There were no statistical significance in ADR between 2 groups(P>0.05). CONCLUSIONS:Sodium hyaluronate eye drops is effective for xerophthalmia,and can relieve the symptoms with good safety.

Objective:Selected virulence factors more than and high virulent Aeromonas hydrophia strain made into inactivated vaccine,to study the immunization effect of inactivated vaccine.Methods:Crucian were vaccinated with formalin-killed vaccine via in-traperitoneal injection.Controls were injected with the same volumes of saline.Then the antibody titres, histopathology and relative percent survival were analyzed from samples of both groups.Results: The antibody in the indirect agglutination reaction could be detected in vaccinated fish once a weeks after immunization and reached highest level 6 weeks after immunization.The histopathology analysis indicated that the vaccine had a good protective effect on crucian target organs.Vaccinated fish showed 100%relative percent survival and the immune period would be 6 month.Conclusion:The vaccine in this study has a significant protective effect on crucian and may be used as effective fish vaccines against bacterial septicemia.

Objective:To study the osseointegration of implant in fresh extraction socket with or without bone grafting.Methods:In the mandibular premolar region of 6 beagle dogs,bone defect in the size of (3~4)mm ×(3~4)mm ×(5 ~6)mm on the mesial wall of the mesial root socket was made.On control side implants were installed immediately into the extraction sockets(group A).On an-other side Bio-Oss grafts and membrane(GBR)were placed following implantation(group B).All animals were sacrificed 3 months af-ter implantation,specimens were examined for histo-morphometric analysis of bone to implant contact and new bone formation.Results:No implant was loosening in the 2 groups.New bone was filled in the bone defect areas in 2 groups.No statistical difference of the per-centage of new bone formation and bone-to-implant contact ration(BIC)was observed between 2 groups.Conclusion:With the defect in a certain size on the root socket wall osseointegration may occur between the new bone and implant without bone transplantation.

ObjectiveTo study the relationship between depression and physical self and related factors of head and neck cancer patients (HNCPs).MethodsOne hundred and twenty-eight HNCPs were assessed with self-rating depression scale,physical self scale,negative physical self scale and symptom checklist-90 (SCL-90).SPSS 16.0 was used to analyze the data.ResultsThe evaluation of appearance,sport characters and figure characters were significantly different between sexual groups,the average physical self scale level of the male patients was higher than that of the females (3.33 ± 0.56 vs.2.87 ± 0.32,P ＜ 0.05).The appearance,sport characters,figure characters and sexual characters of physical self-concept were significantly different between age groups,the body satisfaction scale from higher to lower were under 45 years old patients,beyond 60 years old,between 46 and 59 years old.ConclusionThe evaluation of appearance is reversely correlated with the score of SCL-90,the evaluation of short,appearance and fat in negative physical self are positively correlated with the score of SCL-90.

ObjectiveTo observe the early changes of N-methyl-D-aspartic acid (NMDA) receptor subunit NR1 and NR2B expressions in the hypothalamus so as to discuss the underlying mechanism of the excitability alternation of hypothalamus-pituitary-adrenal (HPA) axis following severe burn in rats.MethodsA total of 40 Sprague-Dawley rats were randomized into the control group and the experimental groups ( including 6,24 and 48 hours after burn) and a 30％ total body surface area full-thickness burn was induced by immersing the shaved dorsum into boiling water for 40 s.Then,the expression changes of the NMDA receptor subunits NR1 and NR2B in the hypothalamus tissue were examined by using immunohistochemistry and Western blot.Results There was an obvious enhancement of the excitability of HPA axis and obvious changes of serum concentration of cortisol and hemodynamics.The serum concentrations of cortisol of the experimental groups reached the peak at 6 hour after burn and were significantly higher than the control group even 48 hours later (P ＜ 0.05 ).Systolic pressure (SP),diastolic pressure (DP) and left ventricular systolic pressure (LVSP) of the experimental groups were all decreased at 6 hours after burn,of which SP and DP were still lower than the control group at 48 hours after burn (P ＜0.05).Based on the Western blot assay,the NR1 level was increased and reached the peak (1.12 ±0.27) in the experimental groups and 0.45 -±0.15 in the control group at 24 hours after burn (P ＜ 0.05).In the meantime,the NR2B level was decreased significantly and reached the lowest for 0.65 ±0.21 at 6 hours after burn,with statistical difference in comparison with 1.38 ±0.51 in the control group (P ＜0.05.The NR2B level recovered to pre-burn level for 1.25 ±0.30 at 48 hours in the experimental groups,with no statistical difference compared with 1.38 ± 0.51 in the control group (P ＞ 0.05 ).The immunohistochemistry results on hypothalamus paraventricular nucleus (PVN) was consistent with the Western blot results.ConclusionThe persistent plasticity changes of hypothalamus subunits NR1 and NR2B may be on of important mechanisms for early excitability alteration of HPA axis after severe burn.

Objective To retrospectively review the efficacy of large craniotomy with subtemporal decompression in the treatment of severe craniocerebral injury combined with cerebral infarction.Methods Forty-eight patients suffering from severe craniocerebral injury combined with cerebral infarction were randomized into the treatment group (managed by large craniotomy decompression) and the control group (managed by clearance of hematoma by routine craniotomy ).A retrospective analysis was carried out to compare the cerebral infarction size between the two groups before operation and at day 1 and 1 week after operation and the outcomes of the two corresponding groups three months post-operatively.Results The infarction size of the treatment group was insignificant in comparison with that of the control group pre-operatively and at day 1 post-operatively,but was significantly smaller than that of the control group one week post-operatively (P ＜ 0.05 ).The mortality and moderate disability/good rate of the treatment and control groups presented significant differences three months post-operatively (21％,42％ vs 33％,25％ respectively,P ＜ 0.05 ).Conclusion Large craniotomy decompression is effective in treating severe craniocerebral injury combined with cerebral infarction and plays an important role in reducing disability and mortality.