Development of an innovative method for the evaluation of fungal contamination of surfaces

The objective of this technical report is to compare the ability to capture fungal spores through samples performed with three different methods: Rodac contact plates, cotton pad and a pad prepared with a dusting cloth (DC pads) selected from those available on the market. The tests were conducted using a suspension of Aspergillus niger conidia equal to 0.5 MacFarland diluted 1/30, 1/40, 1/50, 1/100. With each of these dilutions 3 sterile tiles of stainless steel were contaminated, each divided into 16 small squares, in the center of which 0.1 ml of the dilution chosen was placed and left to dry (for a total of 12 sheets). In addition, we have used other 6 tiles to repeat the experience with dilutions 1/40 and 1/50. A total of 288 squared surfaces were contaminated: 96 of these were sampled with Rodac contact plates, 96 with cotton pads and 96 with DC and then inseminated in Petri plates. Sabouraud dextrose agar was used as culture medium for the first 12 plates, while, for the other 6 plates Sabouraud dextrose agar added with lecithin and polysorbate 80 was used. All plates were incubated at 37 degrees for 18 hours. To estimate the differences among the sampling methods and the dilutions tested, multiple linear regression was used. The analysis showed that the number of colonies harvested at dilution 1/40 is 13% higher (P = 0.09) than the number harvested at dilution 1/50 and the number of colonies harvested at dilution 1/30 is 6% higher than dilution 1/50 (P = 0.52). With regard to the comparison between the number of colonies harvested with Rodac contact plates, with cotton pads and DC pads, regression analysis shows that cotton pads harvest a number of fungal cfu 5 times higher than those detected with Rodac plates, while DC pads harvest a number of fungal ufc 6 times higher than those detected with Rodac plates (P < 0.00005). These results, although preliminary, indicate that DC pads are a sensitive and simple approach for the environmental control of fungal contamination

rf linewidth reduction in a quantum dot passively mode-locked laser subject to external optical feedback

International audienceThe effect of external optical feedback on an InAs/GaAs quantum dot passively mode-locked laser is investigated. The rf linewidth narrows from 8 KHz in the free-running situation to a value as low as 350 Hz under relatively low feedback. The rf linewidth characterization under resonant feedback at a multiple of the laser cavity length validates the prediction of a previous numerical simulation. It is also confirmed that the integrated rms timing jitter varies as the square root of the rf linewidth. The results are promising for the development of compact, monolithic semiconductor mode-locked lasers as low noise optoelectronic oscillators

Optical feedback instabilities in a monolithic InAs/GaAs quantum dot passively mode-locked laser

International audienceThe impact of optical feedback on the direct performance of a monolithic InAs/GaAs quantum dot passively mode-locked laser intended for applications such as multigigahertz interchip/intrachip clock distribution is experimentally investigated. Evaluation of the feedback resistance is an important feature, as the laser is to be monolithically integrated on chip with other devices, in which case optical isolation is difficult. This work shows that a feedback level on the order of −24 dB is detrimental for mode-locking operation, enhancing noise in the rf electrical signal, strongly narrowing the useful mode-locking region as well as causing central frequency shift, and severe instabilities

The Effect of Epstein-Barr Virus Latent Membrane Protein 2 Expression on the Kinetics of Early B Cell Infection

Infection of human B cells with wild-type Epstein-Barr virus (EBV) in vitro leads to activation and proliferation that result in efficient production of lymphoblastoid cell lines (LCLs). Latent Membrane Protein 2 (LMP2) is expressed early after infection and previous research has suggested a possible role in this process. Therefore, we generated recombinant EBV with knockouts of either or both protein isoforms, LMP2A and LMP2B (Δ2A, Δ2B, Δ2A/Δ2B) to study the effect of LMP2 in early B cell infection. Infection of B cells with Δ2A and Δ2A/Δ2B viruses led to a marked decrease in activation and proliferation relative to wild-type (wt) viruses, and resulted in higher percentages of apoptotic B cells. Δ2B virus infection showed activation levels comparable to wt, but fewer numbers of proliferating B cells. Early B cell infection with wt, Δ2A and Δ2B viruses did not result in changes in latent gene expression, with the exception of elevated LMP2B transcript in Δ2A virus infection. Infection with Δ2A and Δ2B viruses did not affect viral latency, determined by changes in LMP1/Zebra expression following BCR stimulation. However, BCR stimulation of Δ2A/Δ2B cells resulted in decreased LMP1 expression, which suggests loss of stability in viral latency. Long-term outgrowth assays revealed that LMP2A, but not LMP2B, is critical for efficient long-term growth of B cells in vitro. The lowest levels of activation, proliferation, and LCL formation were observed when both isoforms were deleted. These results suggest that LMP2A appears to be critical for efficient activation, proliferation and survival of EBV-infected B cells at early times after infection, which impacts the efficient long-term growth of B cells in culture. In contrast, LMP2B did not appear to play a significant role in these processes, and long-term growth of infected B cells was not affected by the absence of this protein. © 2013 Wasil et al

The influence of the accessory genome on bacterial pathogen evolution

Bacterial pathogens exhibit significant variation in their genomic content of virulence factors. This reflects the abundance of strategies pathogens evolved to infect host organisms by suppressing host immunity. Molecular arms-races have been a strong driving force for the evolution of pathogenicity, with pathogens often encoding overlapping or redundant functions, such as type III protein secretion effectors and hosts encoding ever more sophisticated immune systems. The pathogens’ frequent exposure to other microbes, either in their host or in the environment, provides opportunities for the acquisition or interchange of mobile genetic elements. These DNA elements accessorise the core genome and can play major roles in shaping genome structure and altering the complement of virulence factors. Here, we review the different mobile genetic elements focusing on the more recent discoveries and highlighting their role in shaping bacterial pathogen evolution

Construction of a Baculovirus-Silkworm Multigene Expression System and Its Application on Producing Virus-Like Particles

A new baculovirus-silkworm multigene expression system named Bombyx mori MultiBac is developed and described here, by which multiple expression cassettes can be introduced into the Bombyx mori nuclear polyhedrosis virus (BmNPV) genome efficiently. The system consists of three donor vectors (pCTdual, pRADM and pUCDMIG) and an invasive diaminopimelate (DAP) auxotrophic recipient E. coli containing BmNPV-Bacmid (BmBacmid) with a homologous recombination region, an attTn7 site and a loxp site. Two genes carried by pCTdual are firstly inserted into BmBacmid by homologous recombination, while the other eight genes in pRADM and pUCDMIG are introduced into BmBacmid through Tn7 transposition and cre-loxp recombination. Then the invasive and DAP auxotrophic E. coli carrying recombinant BmBacmid is directly injected into silkworm for expressing heterologous genes in larvae or pupae. Three structural genes of rotavirus and three fluorescent genes have been simultaneously expressed in silkworm larvae using our new system, resulting in the formation of virus-like particles (VLPs) of rotavirus and the color change of larvae. The VLPs were purified from hemolymph by ultracentrifugation using CsCl gradients, with a yield of 12.7 µg per larva. For the great capacity of foreign genes and the low cost of feeding silkworm, this high efficient BmMultiBac expression system provides a suitable platform to produce VLPs or protein complexes

Patterning Bacterial Communities on Epithelial Cells

Micropatterning of bacteria using aqueous two phase system (ATPS) enables the localized culture and formation of physically separated bacterial communities on human epithelial cell sheets. This method was used to compare the effects of Escherichia coli strain MG1655 and an isogenic invasive counterpart that expresses the invasin (inv) gene from Yersinia pseudotuberculosis on the underlying epithelial cell layer. Large portions of the cell layer beneath the invasive strain were killed or detached while the non-invasive E. coli had no apparent effect on the epithelial cell layer over a 24 h observation period. In addition, simultaneous testing of the localized effects of three different bacterial species; E. coli MG1655, Shigella boydii KACC 10792 and Pseudomonas sp DSM 50906 on an epithelial cell layer is also demonstrated. The paper further shows the ability to use a bacterial predator, Bdellovibrio bacteriovorus HD 100, to selectively remove the E. coli, S. boydii and P. sp communities from this bacteria-patterned epithelial cell layer. Importantly, predation and removal of the P. Sp was critical for maintaining viability of the underlying epithelial cells. Although this paper focuses on a few specific cell types, the technique should be broadly applicable to understand a variety of bacteria-epithelial cell interactionsopen3

Paradoxical regulation of Bcl-2 family proteins by 17β-oestradiol in human breast cancer cells MCF-7

Tumorigenesis is related to the dysregulation of cell growth or cell death pathways. Hence, elucidation of the mechanisms involved in the modulation of pro- or anti-apoptotic proteins is important in furthering understanding of breast cancer aetiology and may aid in designing prevention and treatment strategies. In the present study, we examined the role of 17β-oestradiol on the regulation of apoptosis in the breast cancer cell line MCF-7. Using multi-probe RNAase protection assays, we found changes in the mRNA levels of several Bcl-2 family proteins upon treatment of MCF-7 cells with 17β-oestradiol. Unexpectedly, we found a paradoxical effects of 17β-oestradiol on two anti-apoptotic proteins Bcl-2 and Bcl-x. Treatment with 17β-oestradiol resulted in up-regulation of Bcl-2 mRNA and protein, but down-regulated Bcl-x(L) mRNA and protein. The effect of 17β-oestradiol on Bcl-x(L) occurred at concentration-dependent fashion. The effect was specific to 17β-oestradiol since other steroid hormones exert no effect on Bcl-x(L). Tamoxifen, an anti-oestrogen, blocked the down-regulation of Bcl-x(L) by 17β-oestradiol demonstrating this effect is oestrogen receptor-dependent. We speculate that different members of the Bcl-2 family proteins may be regulated through different pathway and these pathways may be modulated by 17β-oestradiol. © 1999 Cancer Research Campaig