[Changes of resistant phenotype and CRISPR/Cas system of four Shigella strains passaged for 90 times without antibiotics].

Objective: To explore the stability of resistant phenotypes and changes of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) gene system on four Shigella strains in the absence of antibiotics. Methods: Four clinical isolated Shigella strains that resistant to different antibiotics were consecutive passaged for 90 times without antibiotics. Agar dilution method was used to determine the minimum inhibitory concentration of Shigella strains. After sequence analysis with PCR, CRISPR Finder and Clustal X 2.1 were applied to identify the changes of CRISPR loci in the Shigella strains. Results: After the consecutive transfer of 90 generations, sensitivity to certain antibiotics of four Shigella strains with different drug resistant spectrums increased. Mel-sf1998024/zz resistance to ampicillin, cephalexin, cefotaxime, chloramphenicol decreased, mel-s2014026/sx resistance to norfloxacin, trimethoprim decreased, mel-sf2004004/sx drug resistance to ampicillin, cefuroxime, cefotaxime, chloramphenicol, trimethoprim decreased and mel-sf2013004/bj resistance to chloramphenicol decreased. The spacer of which matched gene codes Cas and its upstream repeat in 3'end of CRISPR3 got lost in mel-sf1998024/zz and mel-sf2013004/bj. Conclusions:Shigella strains could reduce or lose their resistance to some antibiotics after consecutive transfers, without the interference of antibiotics. CRISPR3 locus had dynamic spacers in Shigella strains while CRISPR3 locus and cas genes might have been co-evolved.

The levels of human milk microRNAs and their association with maternal weight characteristics.

BACKGROUND/OBJECTIVES: Specific microRNAs (miRNAs) that are known to have roles in various physiological and pathological processes in mammals are found in human breast milk, but the impact of maternal status on miRNA levels in milk is not fully understood. The objective of this study was to explore maternal and infant factors that can potentially influence the levels of let-7a (MIMAT0000062), miRNA-30B (MIMAT0000420) and miRNA-378 (MIMAT0000732) in human milk, all of which are known to participate in adipogenesis. SUBJECTS/METHODS: Colostrum from 86 mothers and mature milk from 33 mothers were collected. The miRNA levels in these samples were determined using real-time PCR. Correlations between the levels of these miRNAs and lactation duration, maternal weight/BMI before and late in the pregnancy, maternal age, gestational weeks and infant gender were analysed. RESULTS: The levels of let-7a (2.58±0.67) and miRNA-378 (4.64±0.69) in colostrum were higher than those in mature milk (2.39±0.62, 3.62±0.77, P<0.01). Conversely, the level of miRNA-30B was higher in mature milk (4.92±0.57) than in colostrum (4.05±0.61, P<0.01). The levels of miRNA-30B, let-7a and miRNA-378 in colostrum were negatively correlated with maternal pre-pregnancy BMI (P<0.01), and, in mature milk, let-7a was negatively correlated with maternal weight late in the pregnancy (P<0.05). Moreover, miRNA-30B and miRNA-378 were higher in the colostrum received by girls than in that received by boys (P<0.01). This pattern held for miRNA-378 when controlling for maternal pre-pregnancy BMI. CONCLUSIONS: Adipogenesis miRNAs were expressed in both colostrum and mature milk and were related to maternal weight and infant gender. The miRNAs in human milk determined in this study provide a basis for future studies of their biological functions in infants.

IgE+ B cells are scarce, but allergen-specific B cells with a memory phenotype circulate in patients with allergic rhinitis.

BACKGROUND: Despite the critical role of immunoglobulin E (IgE) in allergy, circulating IgE+ B cells are scarce. Here, we describe in patients with allergic rhinitis B cells with a memory phenotype responding to a prototypic aeroallergen. METHODS: Fifteen allergic rhinitis patients with grass pollen allergy and 13 control subjects were examined. Blood mononuclear cells stained with carboxyfluorescein diacetate succinimidyl ester (CFSE) were cultured with Bahia grass pollen. Proliferation and phenotype were assessed by multicolour flow cytometry. RESULTS: In blood of allergic rhinitis patients with high serum IgE to grass pollen, most IgE(hi) cells were CD123+ HLA-DR(-) basophils, with IgE for the major pollen allergen (Pas n 1). Both B and T cells from pollen-allergic donors showed higher proliferation to grass pollen than nonallergic donors (P = 0.002, and 0.010, respectively), whereas responses to vaccine antigens and mitogen did not differ between groups. Allergen-driven B cells that divided rapidly (CD19(mid) CD3(-) CFSE(lo) ) showed higher CD27 (P = 0.008) and lower CD19 (P = 0.004) and CD20 (P = 0.004) expression than B cells that were slow to respond to allergen (CD19(hi) CD3(-) CFSE(mid) ). Moreover, rapidly dividing allergen-driven B cells (CD19(mid) CFSE(lo) CD27(hi) ) showed higher expression of the plasmablast marker CD38 compared with B cells (CD19(hi) CFSE(mid) CD27(lo) ) that were slow to divide. CONCLUSION: Patients with pollen allergy but not control donors have a population of circulating allergen-specific B cells with the phenotype and functional properties of adaptive memory B-cell responses. These cells could provide precursors for allergen-specific IgE production upon allergen re-exposure.

Use of natural antimicrobials to improve the control of Listeria monocytogenes in a cured cooked meat model system.

Concern about nitrite in processed meats has increased consumer demand for natural products manufactured without nitrite or nitrate. Studies on commercial meat products labeled as "Uncured" and "No-Nitrite-or-Nitrate-Added" have shown less control of nitrite in these products and greater potential growth of bacterial pathogens. To improve the safety of the "naturally cured" meats, several natural ingredients were studied in a cured cooked meat model system (80:20 pork, 10% water, 2% salt, and 150 or 50 ppm ingoing sodium nitrite) that closely resembled commercial frankfurters to determine their inhibitory effect on Listeria monocytogenes. Results showed that cranberry powder at 1%, 2% and 3% resulted in 2-4 log cfu/g less growth of L. monocytogenes compared to the control with nitrite alone (P<0.05). Other natural compounds, such as cherry powder, lime powder and grape seed extract, also provided measureable inhibition to L. monocytogenes when combined with cranberry powder (P<0.05).

Comparative observation on effect of electric acupuncture of neiguan (P 6) at chen time versus xu time on left ventricular function in patients with coronary heart disease.

Paired experimental design was adopted in this experiment for comparative observation on effect of electric acupuncture (EA) of Neiguan (P 6) at Chen Time (7 a.m. to 9 a.m.) versus Xu Time (7 p.m. to 9 p.m.) on left ventricular function in patients with coronary heart disease (CHD). The results show that EA performed at Chen Time could improve the left ventricular function of CHD patients as indicated by shortening of PEPI and decrease of PEPI/LVETI ratio; on the contrary, EA performed at Xu Time prolonged PEPI and raised PEPI/LVETI ratio in CHD patients, suggesting impairment of left ventricular function.

[Effect of electro-acupuncture at neiguan P6 on sino-atrial conduction in patients without sick sinus syndrome].

The effects of electro-acupuncture (EA) at Neiguan P6 served as a main point and Jianshi P5 as an adjunct point on sino-atrial conduction were studied in 10 patients with normal sinus nodal function. The results were as follows: During EA and in period of intrinsic heart rate (IHR0) the sinoatrial conduction times (SACT) from sinus node electrogram were significantly shortened than that in control, (P < 0.01 and P < 0.001, respectively). But the SACT could not be further shortened by stimulating the acupoints during IHR0 (P < 0.05). It is suggested that EA at Neiguan P6 as a main point could improve sino-atrial conductivity in most patients with normal sinus node function, which seems to be mediated by autonomic nerve system. The normal value of SACT after autonomic nerve blockade needs to be established.

[Specific cytotoxicity of recombinant interleukin 6-pseudomonas exotoxin fusion protein (IL6-PE40) on LT12 leukemic cells expressing high levels of interleukin 6 receptor (IL6R)].

An acute promyelocytic leukemic cell line LT12 was transduced with IL6R cDNA by electroporation. The resultant LT12-IL6R+ leukemic cells expressed more than 1000 IL6R per cell. The effect of the recombinant IL6-PE40 on both the LT12-IL6R+ cells and the IL6R- parental cells (LT12-IL6R-) was studied by leukemic progenitor colony (CFU-L) formation and 3H-TdR incorporation assays. It was found that IL6-PE40 at concentrations ranging from 1 to 1,000 ng/ml inhibited CFU-L formation and DNA synthesis of LT12-IL6R+ cells in a dose-dependent manner. No inhibition was seen at the same concentration range in LT12-IL6R- cells. At concentrations of 250-1000ng/ml, IL6-PE40 led to only 40% inhibition of DNA synthesis in LT12-IL6R- cells. Inasmuch as rIL6 at the same concentrations had no significant effect on both IL6R+ and IL6R- cells, we concluded it is IL6-PE40 that exerts its highly specific killing on leukemic cells expressing high levels of IL6R.

Sulphate polyanions prolong the incubation period of scrapie-infected hamsters.

The effect of the organic sulphated polyanions, pentosan sulphate (SP54), dextran sulphate 500 (DS500) and suramin, have been tested on golden Syrian hamsters infected with the 263K strain of scrapie by the intraperitoneal (i.p.) or the intracerebral route. SP54 had the greatest effect in prolonging the incubation period of the disease when administered within 2 h of the i.p. inoculum. The same amount of SP54 given 24 h after scrapie inoculation had a potent effect in some animals and no effect in others. This result suggests that SP54 inhibits the uptake of the scrapie agent into the nerve endings and/or carrier cells at the site of the inoculum, i.e. the peritoneum, and that this event occurs in about 24 h. DS500 had a similar although less potent effect (22.4 days delay during the incubation period) than SP54 (54.4 days) when administered within 2 h of scrapie injection by the i.p. route, and suramin had only a minimal effect (10 days). This study suggests that treatment of scrapie and related spongiform encephalopathies of animals and man is possible only before the agent has reached the clinical target areas of the brain.