Phylogenetic diversity of alkaline protease‐producing psychrotrophic bacteria from glacier and cold environments of Lahaul and Spiti,India

The diversity of proteolytic bacteria associated with a glacier and cold environment soils from three different locations in Lahaul and Spiti, India was investigated. Two hundred seventeen bacterial strains were isolated in pure culture. Subsequently these strains were screened for protease‐production and one hundred nine showed protease production. From these protease producing psychrotrophic bacteria twenty showing high enzyme production at low temperature and alkaline pH were characterized and identified. The 16S rRNA phylogenetic analysis revealed that none of the strains showed 100% identity with the validly published species of various genera. Isolates belonged to three classes i.e. Actinobacteria, Gammaproteobacteria and Alphaproteobacteria, and were affiliated with the genera Acinetobacter, Arthrobacter, Mycoplana, Pseudomonas, Pseudoxanthomonas, Serratia and Stenotrophomonas. The optimal growth temperature ranged from 10 to 28 °C and interestingly, high levels of enzyme productions were measured at growth temperatures between 15 and 25 °C, for most of the isolates in plate assay. Most of the isolates were found to produce at least two other hydrolytic enzymes along with protease. The crude protease from one strain was active over broad range of temperature and pH with optima at 30 °C and 7.5, respectively. The protease activity was enhanced by Ca²⁺, dithiothreitol and β‐mercaptoethanol. While Na⁺, Hg²⁺, Zn²⁺, Mn²⁺, phenylmethanesulfonyl fluoride and ethylenediaminetetraacetic acid did not showed much effect on protease activity. The results enrich our knowledge on the psychrotrophic bacterial diversity and biogeographic distribution of enzyme producing bacteria in western Himalaya. (© 2010 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Prokaryotic diversity of a non‐sulfide,low‐salt cold spring sediment of Shawan County,China

The prokaryotic diversity of a non‐sulfide, low‐salt cold spring sediment was investigated by constructing bacterial and archaeal clone libraries of the 16S rRNA gene. 241 bacterial clones were screened, which could be grouped into 86 ribotypes, based on restriction fragment length polymorphism (RFLP) analysis. These were divided into 11 phyla (Actinobacteria, Acidobacteria, Bacteroidetes, Chlorobi, Cyanobacteria, Firmicutes, Gemmatimonadetes, Nitrospirae, Proteobacteria, Planctomycetes and Verrucomicrobia). Of these, Acidobacteria and Proteobacteria were the most dominant, representing 48% and 25% of the total bacteria clone library, respectively. For the archaeal clone library, 121 positive clones were screened and 22 ribotypes were determined. BLAST analysis indicated that all ribotypes were affiliated with the phylum Crenarchaeota. Phylogenetic analysis classified them into three subgroups (Groups I–III). Groups I and III, belonging to the Soil–Freshwater‐subsurface group and Marine group I, respectively, were the dominant groups, representing 50% and 47% of the library, respectively. Of them, 20% of ribotypes were related to the cold‐loving Crenarchaeota. These findings show that bacteria in spring sediments are more diverse than are archaea; in addition, the spring harbors a large number of novel bacterial and archaeal species and maybe exist novel lineages. (© 2010 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Severity of atopic disease inversely correlates with intestinal microbiota diversity and butyrate‐producing bacteria

The reports on atopic diseases and microbiota in early childhood remain contradictory, and both decreased and increased microbiota diversity have been associated with atopic eczema. In this study, the intestinal microbiota signatures associated with the severity of eczema in 6‐month‐old infants were characterized. Further, the changes in intestinal microbiota composition related to the improvement of this disease 3 months later were assessed. The severity of eczema correlated inversely with microbiota diversity (r = ?0.54, P = 0.002) and with the abundance of butyrate‐producing bacteria (r = ?0.52, P = 0.005). During the 3‐month follow‐up, microbiota diversity increased (P < 0.001) and scoring atopic dermatitis values decreased (P < 0.001) in all infants. This decrease coincided with the increase in bacteria related to butyrate‐producing Coprococcus eutactus (r = ?0.59, P = 0.02). In conclusion, the high diversity of microbiota and high abundance of butyrate‐producing bacteria were associated with milder eczema, thus suggesting they have a role in alleviating symptoms of atopic eczema.     

Antibiotic resistance and molecular epidemiology of the beta‐lactamase‐producing Haemophilus influenzae isolated in Chongqing,China

This study aimed to determine the antibiotic resistance and molecular epidemiology of Haemophilus influenzae isolated from children with acute respiratory infection in Chongqing, China. To this end, 1967 H. influenzae isolates from 2006 to 2009 were analysed regarding β‐lactamase production and antibiotic resistance. Ninety‐nine β‐lactamase‐producing H. influenzae isolates from 2010 were analysed for antibiotic resistance and promoter regions of bla_TEM‐1. β‐lactamase production was found in 35.8% (705/1967) of the strains. All ninety‐nine β‐lactamase‐producing strains from 2010 were of the TEM‐1 type as determined by PCR but did not produce the predicted 1075 bp product. According to PCR‐SSCP and DNA sequencing, the promoter regions of bla_TEM‐1 were categorized into 6 genotypes as SSCP1 (Pdel), SSCP2 (Pa/Pb), SSCP3 (P4), SSCP4 (Prpt.b), SSCP5 (2Prpt) and SSCP6 (P3.b). The Pdel, Pa/Pb and Prpt.b were common promoters of bla_TEM‐1 for H. influenzae isolated from children in Chongqing. Strains with Prpt.b were more resistant to ampicillin (AMP) than strains with Pdel, Pa/Pb and P4 (p < 0.05). Therefore, bla_TEM‐1 β‐lactamase is the main mechanism for resistance of H. influenzae to ampicillin in Chongqing. Furthermore, the Prpt.b promoters may be related to the high resistance of H. influenzae to AMP.     

High prevalence of syphilis,HBV, and HCV co‐infection,and low rate of effective vaccination against hepatitis B in HIV‐infected patients in West China hospital

To investigate the epidemiological features and risk factors of HBV, HCV, and syphilis infection among HIV‐infected patients in West China Hospital. A retrospective study was conducted with HIV‐infected patients from 2014 to 2016 in West China hospital, SCU. Serum makers for HBV, HCV, and syphilis were detected. Among 894 HIV‐infected patients, the prevalence of HIV/HBV, HIV/HCV, HIV/syphilis co‐infections was 14.4%, 5.7%, and 18.9% respectively. HIV/HBV/HCV, HIV/HCV/syphilis, and HIV/HBV/syphilis triple co‐infection was 7 (0.7%), 12(1.3%), 29(3.2%) respectively. The rate of effective vaccination against HBV was only 7.7% in HIV‐infected patients. Age (OR = 0.243 95% CI: 0.114 ?0.518), ethnicity (OR = 3.654 95% CI: 1.849‐7.218) and education level (OR = 0.140 95% CI: 0.033‐0.606) are risk factors affecting HIV/HCV co‐infection. A high prevalence of HIV/syphilis, HIV/HBV, and HIV/ HCV co‐infection can be observed in west China. The rate for HIV‐infected patients who were effectively vaccinated against HBV was fewer than 10%.     

Transmission route and introduction of pandemic SARS‐CoV‐2 between China,Italy, and Spain

We present a phylodynamic and phylogeographic analysis of this new severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) virus in this report. A tree of maximum credibility was constructed using the 72 entire genome sequences of this virus, from the three countries (China, Italy, and Spain) available as of 26 March 2020 on the GISAID reference frame. To schematize the current SARS‐CoV‐2 migration scenario between and within the three countries chosen, using the multitype bearth‐death model implemented in BEAST2. Bayesian phylogeographic reconstruction shows that SARS‐CoV‐2 has a rate of evolution of 2.11 × 10^?3 per sites per year (95% highest posterior density: 1.56 × 10^?3 to 3.89 × 10^?3), and a geographic origin in Shanghai, where time until the most recent common ancestor (tMRCA) emerged, according to the analysis of the molecular clock, around 13 November 2019. While for Italy and Spain, there are two tMRCA for each country, which agree with the assumption of several introductions for these countries. That explains also this very short period of subepidermal circulation before the recent events. A total of 8 (median) migration events occurred during this short period, the largest proportion of which (6 events [75%]) occurred from Shanghai (China) to Spain and from Italy to Spain. Such events are marked by speeds of migration that are comparatively lower as compared with that from Shanghai to Italy. Shanghai's R₀ and Italy's are closer to each other, though Spain's is slightly higher. All these results allow us to conclude the need for an automatic system of mixed, molecular and classical epidemiological surveillance, which could play a role in this global surveillance of public health and decision‐making.     

4‐Thiouridine induces dose‐dependent reduction of oedema,leucocyte influx and tumour necrosis factor in lung inflammation

Recent reports demonstrate a role for nucleotides as inflammatory modulators. Uridine, for example, reduces oedema formation and leucocyte infiltration in a Sephadex‐induced lung inflammation model. Tumour necrosis factor (TNF) concentration was also reduced. Previous in vivo observations indicated that 4‐thiouridine might have similar effects on leucocyte infiltration and TNF release. The aim of this study was thus to investigate the effects of 4‐thiouridine in greater detail. We used a Sephadex‐induced acute lung inflammation model in Sprague–Dawley rats. The dextran beads were instilled intratracheally into the lungs, which were excised and examined after 24 h. Sephadex alone led to massive oedema formation and infiltration of macrophages, neutrophils and eosinophils. Microgranulomas with giant cell formations were clearly visible around the partially degraded beads. A significant increase in bronchoalveolar lavage fluid (BALF) content of TNF and leukotrienes was also seen. 4‐Thiouridine co‐administration affected all variables investigated in this model, i.e. oedema, microscopic and macroscopic appearance of lung tissue, total leucocyte and differential leucocyte counts in BALF, TNF and leukotrienes C₄ (LTC₄), LTD₄ and LTE₄ in BALF, indicating a reproducible anti‐inflammatory effect. In conclusion, we have demonstrated that 4‐thiouridine has anti‐inflammatory effects similar to those of uridine. To our knowledge, this is the first demonstration of pharmacological 4‐thiouridine effects in vivo. The results suggest nucleoside/nucleotide involvement in inflammatory processes, warranting further studies on nucleoside analogues as attractive new alternatives in the treatment of inflammatory diseases.     

Allelic and haplotype diversity of HLA‐A,HLA‐B and HLA‐DRB1 gene at high resolution in the Nanning Han population

The distribution of human leucocyte antigen (HLA) allele and haplotype varied among different ethnic populations. In this study, we investigated the allele and haplotype frequencies of HLA‐A, HLA‐B and HLA‐DRB1 loci in the Nanning Han population who live in Guangxi province of China. We identified 26 HLA‐A, 56 HLA‐B and 31 HLA‐DRB1 alleles in 562 Nanning individuals of Han ethnic group by sequence‐based typing method. Of these, the three most common alleles in HLA‐A, HLA‐B and HLA‐DRB1 loci, respectively, were A*11:01 (32.12%), A*02:07 (12.54%), A*24:02 (12.01%); B*46:01 (14.41%), B*15:02 (13.61%), B*40:01 (11.48%); DRB1*15:01 (14.15%), DRB1*16:02 (11.57%) and DRB1*12:02 (10.14%). With the exception of HLA‐DRB1, the p values of the HLA‐A and HLA‐B loci showed that the HLA allelic distribution in this population was in accordance with Hardy–Weinberg expectation (p > 0.05). A total of 173 HLA~A‐B~DRB1 haplotype with a frequency of >0.1% were presented and the three most common haplotype were HLA‐A*33:03~B*58:01~DRB1*03:01 (6.12%), HLA‐A*11:01~B*15:02~DRB1*12:02 (3.39%) and HLA‐A*11:01~B*15:02~DRB1*15:01 (3.22%). The phylogenetic tree and the principal component analysis suggested that Nanning Han population had a relative close genetic relationship with Chinese Zhuang population and a relative distant genetic relationship with Northern Han Chinese. The information will be useful for anthropological studies, for HLA matching in transplantation and disease association studies in the Chinese population.     

Clinical relevance of circulating anti‐ENA and anti‐dsDNA secreting cells from SLE patients and their dependence on STAT‐3 activation

Disturbances of plasma cell homeostasis and auto‐antibody production are hallmarks of systemic lupus erythematosus. The aim of this study was to explore the presence of circulating anti‐ENA and anti‐dsDNA antibody‐secreting cells, to determine their dependence on plasma cell‐niche cytokines and to analyze their clinical value. The study was performed in SLE patients with serum anti‐ENA and/or anti‐dsDNA antibodies (n = 57). Enriched B‐cell fractions and sorted antibody‐secreting cells (CD19^lowCD38^high) were obtained from blood. dsDNA‐ and ENA‐specific antibody‐secreting cells were identified as cells capable of active auto‐antibody production in culture. The addition of a combination of IL‐6, IL‐21, BAFF, APRIL, and CXCL12 to the cultures significantly augmented auto‐antibody production and antibody‐secreting cell proliferation, whereas it diminished apoptosis. The effect on auto‐antibody production was dependent on STAT‐3 activation as it was abrogated in the presence of the JAK/STAT‐3 pathway inhibitors ruxolitinib and stattic. Among patients with serum anti‐dsDNA antibodies, the detection of circulating anti‐dsDNA‐antibody‐secreting cells was associated with higher disease activity markers. In conclusion, auto‐antibody production in response to plasma cell‐niche cytokines that are usually at high levels in SLE patients is dependent on JAK/STAT‐3 activation. Thus, patients with circulating anti‐dsDNA antibody‐secreting cells and active disease could potentially benefit from therapies targeting the JAK/STAT3 pathway.     

Fabrication of Electrospinning Fibers from Spiropyran‐Based Polymeric Nanowires and their Photochromic Properties

Functional nanowires with photochromic spiropyran (SP) species are prepared by reversible addition–fragmentation transfer (RAFT) dispersion polymerization of styrene using poly(4‐vinylpyridine‐co‐spiropyranyl methacrylate) as a macro‐RAFT agent, and then electrospinning technology is used to fabricate fibers from the functional nanowires. The photoisomerization of SP in the nanowires and fibers is studied, and reversible photo­chromism for both nanowires and fibers is observed. Since the merocyanine (ME form) of the SP emits fluorescence and the SP form is non‐fluorescent, the fluorescence of the nanowires and the fibers can be switched on and off upon alternating UV and visible‐light irradiation.

     

The role of cell cycle in retinal development: Cyclin‐dependent kinase inhibitors co‐ordinate cell‐cycle inhibition,cell‐fate determination and differentiation in the developing retina†

The mature retina is formed through multi‐step developmental processes, including eye field specification, optic vesicle evagination, and cell‐fate determination. Co‐ordination of these developmental events with cell‐proliferative activity is essential to achieve formation of proper retinal structure and function. In particular, the molecular and cellular dynamics of the final cell cycle significantly influence the identity that a cell acquires, since cell fate is largely determined at the final cell cycle for the production of postmitotic cells. This review summarizes our current understanding of the cellular mechanisms that underlie the co‐ordination of cell‐cycle and cell‐fate determination, and also describes a molecular role of cyclin‐dependent kinase inhibitors (CDKIs) as co‐ordinators of cell‐cycle arrest, cell‐fate determination and differentiation. Developmental Dynamics 239:727–736, 2010. © 2010 Wiley‐Liss, Inc.     

In vitro synergy of oxacillin and gentamicin against coagulase‐negative staphylococci from blood cultures of neonates with late‐onset sepsis

Coagulase‐negative staphylococci (CoNS) are the leading cause of late‐onset sepsis (LOS) in neonates. Increasing resistance of CoNS to beta‐lactams and aminoglycosides has led to widespread use of vancomycin, which in turn may lead to resistance to vancomycin. Thus, combination therapy of LOS has been advocated. We aimed to determine the interaction of oxacillin and gentamicin against CoNS. In 2005, 34 isolates of oxacillin‐ and gentamicin‐resistant CoNS were obtained from blood samples of neonates with LOS. Combination effect was tested using the checkerboard method, E‐test with the other antibiotic incorporated in the medium (E‐test‐1) and two E‐test strips placed in a cross‐formation (E‐test‐2). Of 34 isolates 61.8%, 53% and 73.5% revealed synergy or an additive effect when tested by the checkerboard method, E‐test‐1 and E‐test‐2, respectively. Results of all three tests were concordant for six (17.6%) isolates, four showing synergy, and two indifference. Our in vitro results support that combination therapy with penicillinase‐resistant penicillin and aminoglycoside can be an alternative to vancomycin.