Cytofluorometric detection of wine lactic acid bacteria: application of malolactic fermentation to the monitoring

In this study we report for the first time a rapid, efficient and cost-effective method for the enumeration of lactic acid bacteria (LAB) in wine. Indeed, up to now, detection of LAB in wine, especially red wine, was not possible. Wines contain debris that cannot be separated from bacteria using flow cytometry (FCM). Furthermore, the dyes tested in previous reports did not allow an efficient staining of bacteria. Using FCM and a combination of BOX/PI dyes, we were able to count bacteria in wines. The study was performed in wine inoculated with Oenococcus oeni (10⁶ CFU ml^?1) stained with either FDA or BOX/PI and analyzed by FCM during the malolactic fermentation (MLF). The analysis show a strong correlation between the numbers of BOX/PI-stained cells determined by FCM and the cell numbers determined by plate counts (red wine: R ² ≥ 0.97, white wine R ² ≥ 0.965). On the other hand, we found that the enumeration of O. oeni labeled with FDA was only possible in white wine (R ² ≥ 0.97). Viable yeast and LAB populations can be rapidly discriminated and quantified in simultaneous malolactic-alcoholic wine fermentations using BOX/PI and scatter parameters in a one single measurement. This rapid procedure is therefore a suitable method for monitoring O. oeni populations during winemaking, offers a detection limit of <10⁴ CFU ml^?1 and can be considered a useful method for investigating the dynamics of microbial growth in wine and applied for microbiological quality control in wineries.     

Synthesis of some pyrazolyl benzenesulfonamide derivatives as dual anti-inflammatory antimicrobial agents

Four series of pyrazolylbenzenesulfonamide derivatives were synthesized and evaluated for their anti-inflammatory activity using cotton pellet induced granuloma and carrageenan-induced rat paw edema bioassays. Moreover, COX-1 and COX-2 inhibitory activity, ulcerogenic effect and acute toxiCIT000y were also determined. Furthermore, the target compounds were screened for their in-vitro antimicrobial activity against Eischerichia coli, Staphylococcus aureus and Candida albicans. Compounds 4-(3-Phenyl-4-cyano-1H-pyrazol-1-yl)benzenesulfonamide 9a and 4-(3-Tolyl-4-cyano-1H-pyrazol-1-yl)benzenesulfonamide 9b were not only found to be the most active dual anti-inflammatory antimicrobial agents in the present study with good safety margin and minimal ulcerogenic effect but also exhibited good selective inhibitory activity towards COX-2. A docking pose for 9a and 9b separately in the active site of the human COX-2 enzyme was also obtained. Therefore, these compounds would represent a fruitful matrix for the development of dual anti-inflammatory antimicrobial candidates with remarkable COX-2 selectivity.     

Primary cilium depletion typifies cutaneous melanoma in situ and malignant melanoma

Cutaneous melanoma is a lethal malignancy that arises spontaneously or via in situ precursor neoplasms. While melanoma in situ and locally invasive malignant melanoma can be cured surgically, these lesions can sometimes be difficult to distinguish from melanocytic nevi. Thus, the identification of histolopathologic or molecular features that distinguish these biologically distinct lesions would represent an important advance. To this end, we determined the abundance of melanocytic primary cilia in a series of 62 cases composed of typical cutaneous melanocytic nevi, melanoma in situ, invasive melanoma, and metastatic melanoma. Primary cilia are sensory organelles that modulate developmental and adaptive signaling and notably, are substantially depleted from the neoplastic epithelium of pancreatic carcinoma at a stage equivalent to melanoma in situ. In this series, we find that while nearly all melanocytes in 22 melanocytic nevi possessed a primary cilium, a near-complete loss of this organelle was observed in 16 cases of melanoma in situ, in 16 unequivocal primary invasive melanomas, and in 8 metastatic tumors, each associated with a cutaneous primary lesion. These findings suggest that the primary cilium may be used to segregate cutaneous invasive melanoma and melanoma in situ from melanocytic nevi. Moreover, they place the loss of an organelle known to regulate oncogenic signaling at an early stage of melanoma development.     

Growth establishment and phenology of four woody Saharan species

Henophyton deserti, Calligonum polygonoides, Ephedra alata and Anthyllis henoniana may be suitable Saharan species for the rehabilitation of degraded lands. In this context, the phenology, excavation of root systems and seedling establishment of these species were investigated in the southern Tunisia. Our data suggest that phenological differences between the studied species may depend on the rooting depth. The mean seedling emergence rate was 73% in H. deserti, 66% in A. henoniana, 60% in C. polygonoides and 50% in E. alata. Statistical analyses of seedling establishment showed a highly significant difference among the four species in both seasons (F = 1595.15, P < 0.0001), species (F = 781.57, P < 0.0001) and species/season interaction (F = 61.84, P < 0.0001). After 3 years, seedling mortality ranged from 73% to 89%. Seedling mortality reached a maximum in the summer of the first year of study (P < 0.001). This study provided further understanding of the ecological tolerance, adaptive strategy and the low density of these species in nature. Henophyton deserti appears to be better adapted to the harsh desert environment. These findings explain the very high density observed in the Sahara compared with other studied species. This species has therefore proven to be an excellent candidate for the ecological restoration of degraded desert rangelands and can be introduced successfully in to rehabilitation programmes.     

Investigation of Alteration in the Levels of Iron and Copper in Scalp Hair Samples of Patients Having Different Types of Viral Hepatitis

The aim of this study was to measure the alterations of copper and iron contents in scalp hair samples of hepatitis A–E patients of both genders, same age group, and socioeconomic status. For comparative study, the scalp hair samples of healthy individuals of the same age and socioeconomic status were collected. The concentrations of copper and iron were measured using an atomic absorption spectrophotometer, after microwave-assisted acid digestion. The validity and accuracy of methodology was checked using a certified reference material. The results of this study showed that the mean values of copper and iron were higher in scalp hair samples of hepatitis patients than those of age-matched control subjects, while the difference was significant in the cases of patients having viral hepatitis B, C, and D as compared to those who have viral hepatitis A and E (p?<?0.001). It was concluded that the overload of copper and iron in the human body may cause lipid peroxidation and eventually damage the hepatic system.     

Determination of the CCR5∆32 frequency in Emiratis and Tunisians and the screening of the CCR5 gene for novel alleles in Emiratis

Background

The chemokine receptor components play crucial roles in the immune system and some of them serve as co-receptors for the HIV virus. Several studies have documented that variants in chemokine receptors are correlated with susceptibility and resistance to infection with HIV virus. For example, mutations in the chemokine receptor 5 gene (CCR5) resulting in loss-of-function (such as the homozygous CCR5?32) confer high degree of resistance to HIV infection. Heterozygotes for these variants exhibit slow progression to AIDS. The prevalence of CCR5 polymorphisms varies among ethnic and geographical groups. For example, the CCR5?32 variant is present in 10–15% of north Europeans but is rarely encountered among Africans. This study aims to identify the prevalence of some CCR5 variants in two geographically distant Arab populations (namely Emiratis and Tunisians).

Methodology

The prevalence of CCR5 gene variants including CCR5?32, FS299, C101X, A29S and C178R has been determined using PCR and direct DNA sequencing. A total of 403 unrelated healthy individuals (253 Emiratis and 150 Tunisians) were genotyped for the CCR5?32 variant using PCR amplification and gel electrophoresis. In addition, 200 Emiratis have been screened for other SNPs using Sanger DNA sequencing.

Results

Among Emiratis, the allele frequency of the CCR5?32 variant has been found to be 0.002. In addition, two variants L55Q and A159 were found at a frequency of 0.002. Moreover, the prevalence of the CCR5?32 variant in Tunisians was estimated to be 0.013 which is relatively higher than its frequency in Emiratis but lower than Europeans.

Conclusion

We conclude that the allele frequency of the most critical CCR5 polymorphism (?32) is extremely low among Emiratis compared to other Arabs and North Europeans. In addition, very low allele frequencies of other CCR5 polymorphisms have been detected among Emiratis.     

Resting State Networks' Corticotopy: The Dual Intertwined Rings Architecture

How does the brain integrate multiple sources of information to support normal sensorimotor and cognitive functions? To investigate this question we present an overall brain architecture (called “the dual intertwined rings architecture”) that relates the functional specialization of cortical networks to their spatial distribution over the cerebral cortex (or “corticotopy”). Recent results suggest that the resting state networks (RSNs) are organized into two large families: 1) a sensorimotor family that includes visual, somatic, and auditory areas and 2) a large association family that comprises parietal, temporal, and frontal regions and also includes the default mode network. We used two large databases of resting state fMRI data, from which we extracted 32 robust RSNs. We estimated: (1) the RSN functional roles by using a projection of the results on task based networks (TBNs) as referenced in large databases of fMRI activation studies; and (2) relationship of the RSNs with the Brodmann Areas. In both classifications, the 32 RSNs are organized into a remarkable architecture of two intertwined rings per hemisphere and so four rings linked by homotopic connections. The first ring forms a continuous ensemble and includes visual, somatic, and auditory cortices, with interspersed bimodal cortices (auditory-visual, visual-somatic and auditory-somatic, abbreviated as VSA ring). The second ring integrates distant parietal, temporal and frontal regions (PTF ring) through a network of association fiber tracts which closes the ring anatomically and ensures a functional continuity within the ring. The PTF ring relates association cortices specialized in attention, language and working memory, to the networks involved in motivation and biological regulation and rhythms. This “dual intertwined architecture” suggests a dual integrative process: the VSA ring performs fast real-time multimodal integration of sensorimotor information whereas the PTF ring performs multi-temporal integration (i.e., relates past, present, and future representations at different temporal scales).     

Characterization of the Viable but Nonculturable (VBNC) State in Saccharomyces cerevisiae

The Viable But Non Culturable (VBNC) state has been thoroughly studied in bacteria. In contrast, it has received much less attention in other microorganisms. However, it has been suggested that various yeast species occurring in wine may enter in VBNC following sulfite stress.In order to provide conclusive evidences for the existence of a VBNC state in yeast, the ability of Saccharomyces cerevisiae to enter into a VBNC state by applying sulfite stress was investigated. Viable populations were monitored by flow cytometry while culturable populations were followed by plating on culture medium. Twenty-four hours after the application of the stress, the comparison between the culturable population and the viable population demonstrated the presence of viable cells that were non culturable. In addition, removal of the stress by increasing the pH of the medium at different time intervals into the VBNC state allowed the VBNC S. cerevisiae cells to “resuscitate”. The similarity between the cell cycle profiles of VBNC cells and cells exiting the VBNC state together with the generation rate of cells exiting VBNC state demonstrated the absence of cellular multiplication during the exit from the VBNC state. This provides evidence of a true VBNC state. To get further insight into the molecular mechanism pertaining to the VBNC state, we studied the involvement of the SSU1 gene, encoding a sulfite pump in S. cerevisiae. The physiological behavior of wild-type S. cerevisiae was compared to those of a recombinant strain overexpressing SSU1 and null Δssu1 mutant. Our results demonstrated that the SSU1 gene is only implicated in the first stages of sulfite resistance but not per se in the VBNC phenotype. Our study clearly demonstrated the existence of an SO₂-induced VBNC state in S. cerevisiae and that the stress removal allows the “resuscitation” of VBNC cells during the VBNC state.