The archetype Pseudomonas aeruginosa proteins TssB and TagJ form a novel subcomplex in the bacterial type VI secretion system

In Pseudomonas aeruginosa three type VI secretion systems (T6SSs) coexist, called H1‐ to H3‐T6SSs. Several T6SS components are proposed to be part of a macromolecular complex resembling the bacteriophage tail. The T6SS protein HsiE1 (TagJ) is unique to the H1‐T6SS and absent from the H2‐ and H3‐T6SSs. We demonstrate that HsiE1 interacts with a predicted N‐terminal α‐helix in HsiB1 (TssB) thus forming a novel subcomplex of the T6SS. HsiB1 is homologous to the Vibrio cholerae VipA component, which contributes to the formation of a bacteriophage tail sheath‐like structure. We show that the interaction between HsiE1 and HsiB1 is specific and does not occur between HsiE1 and HsiB2. Proteins of the TssB family encoded in T6SS clusters lacking a gene encoding a TagJ‐like component are often devoid of the predicted N‐terminal helical region, which suggests co‐evolution. We observe that a synthetic peptide corresponding to the N‐terminal 20 amino acids of HsiB1 interacts with purified HsiE1 protein. This interaction is a common feature to other bacterial T6SSs that display a TagJ homologue as shown here with Serratia marcescens. We further show that hsiE1 is a non‐essential gene for the T6SS and suggest that HsiE1 may modulate incorporation of HsiB1 into the T6SS.     

Seasonal variations of cellular stress response in the heart and gastrocnemius muscle of the water frog (Pelophylax ridibundus)

The present study aimed to investigate the seasonal cellular stress response in the heart and the gastrocnemius muscle of the amphibian Pelophylax ridibundus (former name Rana ridibunda) during an 8 month acclimatization period in the field. Processes studied included heat shock protein expression and protein kinase activation. The cellular stress response was addressed through the expression of Hsp70 and Hsp90 and the phosphorylation of stress-activated protein kinases and particularly p38 mitogen-activated protein kinase (p38 MAPK), the extracellular signal-regulated kinases (ERK-1/2) and c-Jun N-terminal kinases (JNK1/2/3). Due to a general metabolic depression during winter hibernation, the induction of Hsp70 and Hsp90 and the phosphorylation of p38 MAPK, JNKs and ERKs are retained at low levels of expression in the examined tissues of P. ridibundus. Recovery from hibernation induces increased levels of the specific proteins, probably providing stamina to the animals during their arousal.     

Anti-tuberculosis therapy-induced hepatotoxicity among Ethiopian HIV-positive and negative patients

Background

To assess and compare the prevalence, severity and prognosis of anti-TB drug induced hepatotoxicity (DIH) in HIV positive and HIV negative tuberculosis (TB) patients in Ethiopia.

Methodology/Principal Findings

In this study, 103 HIV positive and 94 HIV negative TB patients were enrolled. All patients were evaluated for different risk factors and monitored biochemically and clinically for development of DIH. Sub-clinical hepatotoxicity was observed in 17.3% of the patients and 8 out of the 197 (4.1%) developed clinical hepatotoxicity. Seven of the 8 were HIV positive and 2 were positive for HBsAg.

Conclusions/Significance

Sub-clinical hepatotoxicity was significantly associated with HIV co-infection (p = 0.002), concomitant drug intake (p = 0.008), and decrease in CD4 count (p = 0.001). Stepwise restarting of anti TB treatment was also successful in almost all the patients who developed clinical DIH. We therefore conclude that anti-TB DIH is a major problem in HIV-associated TB with a decline in immune status and that there is a need for a regular biochemical and clinical follow up for those patients who are at risk.     

Electrical Stimulation Improves Microbial Salinity Resistance and Organofluorine Removal in Bioelectrochemical Systems

Fed batch bioelectrochemical systems (BESs) based on electrical stimulation were used to treat p-fluoronitrobenzene (p-FNB) wastewater at high salinities. At a NaCl concentration of 40 g/liter, p-FNB was removed 100% in 96 h in the BES, whereas in the biotic control (BC) (absence of current), p-FNB removal was only 10%. By increasing NaCl concentrations from 0 g/liter to 40 g/liter, defluorination efficiency decreased around 40% in the BES, and in the BC it was completely ceased. p-FNB was mineralized by 30% in the BES and hardly in the BC. Microorganisms were able to store 3.8 and 0.7 times more K⁺ and Na⁺ intracellularly in the BES than in the BC. Following the same trend, the ratio of protein to soluble polysaccharide increased from 3.1 to 7.8 as the NaCl increased from 0 to 40 g/liter. Both trends raise speculation that an electrical stimulation drives microbial preference toward K⁺ and protein accumulation to tolerate salinity. These findings are in accordance with an enrichment of halophilic organisms in the BES. Halobacterium dominated in the BES by 56.8% at a NaCl concentration of 40 g/liter, while its abundance was found as low as 17.5% in the BC. These findings propose a new method of electrical stimulation to improve microbial salinity resistance.     

Radiochemical and radiobiological assessment of a pyridyl-S-cysteine functionalized bombesin derivative labeled with the 99mTc(CO)3+ core

Bombesin is a neuropeptide widely studied due to its ability to target various types of cancers. Technetium-99m on the other hand is ideal for diagnostic tumor targeting. The aim of the present study is the investigation of the coupling of the ligand (S)-(2-(2′-pyridyl)ethyl)-d,l-cysteine with the BN-peptide Gln-Arg-Leu-Gly-Asn-Gln-Trp-Ala-Val-Gly-His-Leu-Met(CONH₂) through the spacer aminohexanoic acidand the labeling of the resulting derivative MBN with the synthon [M(CO)₃(H₂O)₃]⁺ (M = ^99mTc, Re). The peptide was synthesized according to the SPPS method, purified and characterized by ESI-MS. The new ^99mTc-labeled biomolecule was stable in vitro, showed high affinity for the human GRP receptor expressed in PC3 cells and the rate of internalization was found to be time-dependent tissue distribution of the radiopeptide was evaluated in normal mice and in prostate cancer experimental models and significant radioactivity uptake was observed in the pancreas of normal mice as well as in PC3 tumors. Dynamic studies of the radiopeptide showed satisfactory tumor images.     

Effect of temperature on development, overwintering and establishment potential of Franklinothrips vespiformis in the UK

This study investigated the effect of temperature on the development and overwintering potential of the predatory thrips Franklinothrips vespiformis (Crawford) (Thysanoptera: Aeolothripidae), a biological control agent used against glasshouse pests in continental Europe and Israel. Developmental rates increased linearly with rearing temperatures. It was estimated that 304.9 degree days, above a lower threshold temperature of 11.9 °C, were required for F. vespiformis to complete development from egg to adult eclosion. The effect of low temperatures (–5, 0, and 5 °C) was examined on adult female and larval survival. Subsequent reproductive and developmental attributes of survivors were also investigated. Lethal time experiments indicated that larval stages are more cold tolerant than adult F. vespiformis females. Surviving larvae increased their developmental times to adults with decreasing temperature and increasing exposure periods and second instars were significantly more successful than first instars in reaching adulthood. Surviving adult females decreased their oviposition rate with decreasing temperature and increasing exposure periods, and exposures to low temperatures affected the number of viable eggs produced. The results are discussed in the context of overwintering and establishment potential of F. vespiformis in the UK in the event of introducing the predatory thrips as a biological control agent against glasshouse pests.     

Evaluation of the role of the vaccinia virus uracil DNA glycosylase and A20 proteins as intrinsic components of the DNA polymerase holoenzyme

The vaccinia virus DNA polymerase is inherently distributive but acquires processivity by associating with a heterodimeric processivity factor comprised of the viral A20 and D4 proteins. D4 is also an enzymatically active uracil DNA glycosylase (UDG). The presence of an active repair protein as an essential component of the polymerase holoenzyme is a unique feature of the replication machinery. We have shown previously that the A20-UDG complex has a stoichiometry of ～1:1, and our data suggest that A20 serves as a bridge between polymerase and UDG. Here we show that conserved hydrophobic residues in the N' terminus of A20 are important for its binding to UDG. Our data argue against the assembly of D4 into higher order multimers, suggesting that the processivity factor does not form a toroidal ring around the DNA. Instead, we hypothesize that the intrinsic, processive DNA scanning activity of UDG tethers the holoenzyme to the DNA template. The inclusion of UDG as an essential holoenzyme component suggests that replication and base excision repair may be coupled. Here we show that the DNA polymerase can utilize dUTP as a substrate in vitro. Moreover, uracil moieties incorporated into the nascent strand during holoenzyme-mediated DNA synthesis can be excised by the viral UDG present within this holoenzyme, leaving abasic sites. Finally, we show that the polymerase stalls upon encountering an abasic site in the template strand, indicating that, like many replicative polymerases, the poxviral holoenzyme cannot perform translesion synthesis across an abasic site.     

The Effect of Farmer Attitudes and Objectives on the Heterogeneity of Farm Attributes and Management in Switzerland

Identifying and interpreting the heterogeneity of farmer behaviour is becoming increasingly important in support of policy- and decision-making goals. This paper explores whether observed differences in farming practices can be interpreted from the heterogeneity of farmer behaviour. Farmer attitudes and objectives were analysed using a combination of principal components and cluster analysis applied to responses to statements in a telephone-based survey. Respondents were classified into four profiles; business-oriented, lifestylers, multifunctionalists and traditionalists. Each profile differed in terms of farm management practices, the amount of land farmers either managed or owned, the existence of successors and the importance placed on household members in providing information. The results suggest that knowledge of farmer behavioural profiles could support more targeted policy development that accounts for alternative farmer goals. However, similarities were also found between the profiles, suggesting that farmer behaviour would be better interpreted as a dynamic set of identities, rather than as static profiles.     

Macro-botanical evidence for plant use at Neolithic Çatalhöyük south-central Anatolia, Turkey

Analysis of charred plant macro-remains, including wood charcoals, cereals, seeds, tubers and fruits from the Neolithic site of ?atalh?yük has indicated complex patterns of plant resource use and exploitation in the Konya plain during the early Holocene. Evidence presented in this paper shows that settlement location was not dictated by proximity to high quality arable land and direct access to arboreal resources (firewood, timber, fruit producing species). A summary of the patterns observed in sample composition and species representation is outlined here together with preliminary interpretations of these results within their broader regional context. Received September 4, 2001 / Accepted April 9, 2002