Upstream Box Research Articles

Hydrodynamics of fluid resonance in a narrow gap between two boxes with different breadths

Violent fluid oscillations occur inside narrow gaps between closely side-by-side structures, threating the safety of the operations. In the current study, fluid resonance in the gap between two side-by-side different boxes is investigated by a two-dimensional viscous-flow numerical wave tank, which can simultaneously predict the viscous damping and free-surface nonlinearity. The accuracy of the established numerical model is fore validated by the published experimental data. The upstream box keeps fixed and has six varied breadths, the downstream box heaves freely under wave actions and remains a constant breadth. The current work aims to study the effects of the breadth ratio and heave motion of the lee-side box on the hydrodynamics of gap resonance. The wave amplification in the gap, the heave motions of the lee-side box, the reflection, transmission, and energy loss coefficients, and the response time and damping time are concerned. Results show that the gap resonance frequency is insensitive to the variation of the breadth ratio. As the breadth ratio increases from 0.4 to 1.4. The maximum wave height amplification in the gap reduces more than 25%. The maximum response time of the fluid resonance occurs when two boxes have equal breadth.

Experimental Investigation about the Effects of Blockage on Upstream Box Culverts

This experimental study focuses on scouring in box culvert inlets under steady-state conditions and at different percentages of blockage ranging from 0% to 65%, and also looks at the hydraulics of water in the culvert. The investigation shows that the blockage of the culverts has influenced the scouring pattern at the blocked culverts' entrances. Ten experiments were carried out at the laboratory to see how blockage impacts the scouring pattern upstream of a box culvert during steady flow. Both partially blocked and unblocked cases were implemented in this study. The experimental tests were done until the equilibrium scour occurred, which took about 3.5 hours of water flow to reach equilibrium conditions. The results revealed that the blockage will increase the water depth at the inlet by about 30%–50%, which may affect the safety of structures or cause culvert failure. In addition, the results discovered that the maximum scour depth, which inversely correlated with the obstruction upstream of the box culvert, increased with increasing discharge.

Aerodynamic force distribution and vortex drifting pattern around a double-slotted box girder under vertical vortex-induced vibration

Double-slotted box girder is a novel type of main girder for long-span bridges. To investigate the vertical vortex induced vibration (VIV) performance and aerodynamic effect of double-slotted box girder, wind tunnel tests were conducted on a spring-suspended sectional model. The results showed that the aerodynamic forces on the upper and lower surfaces of upstream box, the upper surface of middle box, and the rear regions of upper and lower surfaces of downstream box contribute positively to the VIV, becoming the main excitation source. Compared with streamlined closed-box and central-slotted box girders, the regions with obvious contribution values on double-slotted box girder surface significantly increase. This indicates that double slots allow the aerodynamic forces acting on larger areas of girder surface to participate in VEF generation and potentially affect the VIV performance. A vortex-related flow pattern around the double-slotted box girder was described by combining the vortex drifting theory and simplified vortex model, which provide insight into the mechanism of vertical VIV. Periodic vortex drifting exists on the upper surface of downstream box. The aerodynamic forces from the windward side to the front area of the lower surface of middle box appear to be significantly affected by vortices in windward slot.

Investigation of Blockage Upstream Box Coverage on the Scour Pattern in the Open Channel

Under sub-critical flow conditions, the presence of blockage (solid wastes and weeds) upstream of box coverage and the scours pattern downstream of the coverage was explored in this study. The upstream blocking was simulated using 48 runs in an artificial trapezoidal cross-section with three square box coverages of side dimensions 8.80, 10.40, and 12.90 cm, four water flows of 2, 5, 8, and 11 L/s, and four blockage ratios of 0, 10, 20, and 30% relative to the coverage cross-section area. To estimate the scour hole characteristics, a 2.00 m long, 0.60 m bed width, and 0.30 m deep sand basin filled with D50 = 0.50 mm bed material was constructed directly downstream of the coverage outlet. In each run, the water level, velocity, and scoured hole parameters downstream of the coverage were measured. According to the analysis, the Non-blocked coverage has less scour depth and length than the partially blocked coverage, where the maximum scour depths and length of the Non- blocked coverage for cases 1, 2, and 3 at the discharge of 11 l/s were 73, 72, and 17 % respectively, and 77.56, 77.34, and 83.66 % respectively relative to the maximum scour depth and length of partially blocked coverage at the discharge of 11 l/s and blockage ratio 30 %. The depth and length of the scour hole downstream coverage are increased with the increment of the coverage's blockage ratio and discharge and are reversely proportional to the inlet area of coverage. Increased relative scour depth and length by 3.60 percent and 11.80 percent respectively, by increasing the downstream Froude number of flow (Frd) by 0.01 while the coverage’ relative wetted area (Ar) is constant. The study suggested applying coverage with a suitable area and water discharge, and protection techniques downstream of the coverage to reduce the influence of coverage and blockage on the open channel's hydraulic efficiency and the scour pattern downstream of the coverage. Also, install a trash rack upstream of the covering and remove aquatic weeds and solid wastes periodically upstream and inside the coverage.

Analysis of The Impact of Sedimentation Dredging on Headloss Repair & Operating Patterns of CWP (Circulating Water Pump) PLTU Lontar

The PLTU Lontar Extension Project (1x315 MW) is a national strategic project being built by PT. PLN (Persero). The project stage is currently in the commissioning phase but is hampered by indications of sedimentation occurring at the intake mouth area of ??the Circulating Water System. There are indications of sedimentation at the intake mouth due to the high head loss between the intake mouth and the upstream box culvert which has an impact on inhibiting the flow of seawater entering the Lontar PLTU intake canal. The existence of obstacles to the flow of sea water has resulted in insufficient cooling water flow for the needs of 8 Circulating Water Pump (CWP) pumps, namely 6 CWP for 3 units of PLTU Existing and 2 CWP for 1 unit of PLTU Extension. The sea bed level on the side of the intake mouth according to the basic design is -4.5 m but after the survey (bathymetry, topography, floating sediment, currents, tides) the actual condition is currently at a depth of -0.9 m. As one solution, the dredging method is taken to restore conditions according to the basic design. To find out if there is an improvement in headloss, a simulation is carried out using the Hec-ras 2D application. After the simulation, it was found that the headloss/compression loss significantly improved in the hydraulic simulation with a flow rate of 42 m3/s at 6 pumps running, before dredging 30 cm and after dredging to 18 cm and also with a flowrate of 60 m3/s at 8 pumps running. , before dredging 55 cm and after dredging to 21 cm at the right position upstream of the box culvert. This headloss condition is predicted to propagate downstream to the forebay and CWP pump pits, so that dredging in this area is simulated to increase the performance of the Lontar PLTU circulating water intake so that 8 CWPs can operate together continuously.

Vortex-excited force evolutionary characteristics of split three-box girder bridges during vortex-induced vibration

As novel cross sections, aerodynamic behaviors and vortex-induced vibration (VIV) inducement of split box girders, especially split three-box girders, are not fully understood. Analysis of vortex-excited force (VEF) evolutionary characteristics in the VIV process may reveal valuable information. Based on displacement and pressure measurement wind tunnel tests on a large-scale section model, the evolutionary characteristics of typical VEF indexes were expounded. VIV was divided into four stages: pre-VIV, ascent, extreme point, and descent. At the extreme point, high-order harmonics were found, which indicates that VEF has obvious nonlinearities. Mean and RMS values of global VEF appear to be controlled by upstream and downstream boxes. An increase in RMS values is the main cause of VIV. VEF spanwise correlation is mainly dominated by structural vibration at the extreme point, while spanwise distance is dominant in other stages. Local aerodynamic forces on the surfaces include upstream box surface (excluding partial lower surfaces of the wind fairing), upstream and downstream curved surfaces in upstream gap, upper surface of downstream box, and the wake are the main contributors to global VEF and the main cause of VIV. This work provides a comprehensive understanding of aerodynamic behaviors and VIV inducement of split three-box girder bridges.

Investigation on aerodynamic force nonlinear evolution for a central-slotted box girder under torsional vortex-induced vibration

Central-slotted box girders are widely employed in long-span bridges owing to their advantageous flutter stability. However, the existence of a central slot can degrade their performance under vortex-induced vibrations (VIVs). To clarify the aerodynamic characteristics of a typical central-slotted box girder during VIVs, sectional model wind tunnel tests involving the synchronous measurement of pressure distributions and VIV responses were performed. The computational fluid dynamics (CFD) technique was also used to demonstrate the flow pattern development during VIV before qualitatively explaining the VIV mechanism of the central-slotted box girder. The surface pressure distributions at various amplitude-dependent VIV stages were measured and examined. The evolution of the aerodynamics was investigated from the perspective of the work done by the vortex-excited force (VEF). It was found that the aerodynamic effects on a central-slotted box girder during VIVs are featured with apparent nonlinear evolutionary characteristics. During the lock-in period, the wind-induced pressures at both upper and lower surfaces of the downstream box and the pressures at upper surface of the upstream box make greater contributions to the VEF, which are the main excitation sources of the torsional VIV. However, the contributions of the downstream and upstream boxes are opposite, showing positive and negative correlations with the VEF, respectively. It demonstrated that the positive contribution of the pressures at both upper and lower surfaces of the downstream box weakens the VIV performance of the central-slotted box girder as compared with a streamlined box girder. In addition, the central slot, which improves the correlating flow between the upper and lower regions around the downstream box, causes the distributed aerodynamic forces in these regions to enhance the VEF and perform positive work. This provides a reasonable explanation as to why the VIV effects of central-slotted box girders are typically stronger than those of streamlined box girders.

Vortex-induced Vibration and Control of Split Three-Box Girder Bridges

The vortex-induced vibration (VIV) performance and aerodynamic mechanism of VIV control of a split three-box girder bridge were investigated numerically by Computational Fluid Dynamics (CFD). Large-scale section model wind tunnel tests show that the integrated countermeasure combined by 30% ventilation ratio grids and new scheme for maintenance tracks can effectively restrain vertical and torsional VIVs. The reliability of numerical method is verified by comparing static three-component coefficients and Strouhal numbers obtained from CFD and wind tunnel tests. The flow structure and changes of aerodynamic forces were analyzed via delayed detached-eddy simulation (DDES). For the original section, large-scale vortices form with shedding behind the safety barrier of the upstream box and behind the maintenance tracks, as well above the downstream highway box, introducing periodic pulsating lifts and moments that result in severe VIVs. The grids can effectively reduce the size of vortices in the gaps, prevent the flow from fluctuating and reduce the pressure difference between upper and lower surfaces. The torsional VIVs at non-negative attack angles are not controlled because the grids cannot interfere with the vortices behind the maintenance tracks. The new scheme for maintenance tracks, which has grid-like effects, can eliminate the torsional VIVs at non-negative attack angles by preventing the vortices behind maintenance tracks from impacting downstream boxes. The aerodynamic force components are analyzed to find that box 1 mainly contributes the mean values and box 3 mainly contributes the fluctuating values. The integrated countermeasure can reduce the mean and RMS values of aerodynamic forces to varying degrees. The streamlines and RMS pressure contours indicate that the integrated countermeasure effectively reduced the wake height and vortices motion range, which is conducive to VIV control. This work lays a solid foundation for an in-depth understanding of the aerodynamic characteristics and optimization mechanism of split three-box girder bridges.

Effects of free heave motion on wave resonance inside a narrow gap between two boxes under wave actions

Fluid resonance inside a narrow gap between two side-by-side boxes is investigated based on an open-source CFD package, OpenFOAM. An upstream box heaves freely under wave actions and a downstream box remains fixed. The focus of this work is to study the influence of the motion of the upstream box on the hydrodynamic behavior of the resonant fluid inside the gap. The hydrodynamic behavior considered in this study includes the wave height inside the gap, heave displacement and their harmonic components, and reflection, transmission and energy loss coefficients. For comparison, the configuration in which the two boxes are fixed is considered. It was found that the heave motion of the upstream box increases the fluid resonant frequency and significantly reduces the resonant wave height in the gap. The frequencies at which the maximum and minimum heave displacements of the upstream box are observed to obviously deviate from the fluid resonant frequency. For the wave height in the gap and heave displacement, the effects of the incident wave height on their harmonic components are different. The heave motion of the upstream box results in a larger reflection coefficient and smaller energy loss coefficient.

Influence of gap width on buffeting force spatial correlation and aerodynamic admittance of twin-box bridge deck

Wind tunnel tests, including pressure measurement and Particle Image Velocimetry (PIV) test, have been carried out on twin-box bridge decks with four different gap widths (two small gap widths and two large gap widths). The effect of gap width on the buffeting force coherence and aerodynamic admittance of twin-box bridge deck are presented and discussed. The mechanisms of related experimental result are discussed in detail based on the pressure distributions and flow structures obtained in the tests. The measurements reveal that under the influence of vortices shed from the rear edge of the upstream box, the spanwise coherence of buffeting force on the downstream box decrease significantly with the increase of gap width, leading a sudden reduction in total buffeting force coherence on the twin-box bridge deck. While the aerodynamic admittance of twin-box girder increases conspicuously with the increase of gap width due to the generation of the local flow separation at the leading edge of downstream box for large gap width cases.

Investigation of flutter performance of a twin-box bridge girder at large angles of attack

Twin-box bridge girders are considered to be aerodynamically favourable, and this paper presents a study on the flutter performance of this type of girders at large angles of attack by both wind tunnel tests and computational fluid dynamics (CFD) simulations. The stationary aerodynamic performance of a twin-box bridge girder at different angles of attack is studied, and different central slot widths are considered as well. Then, the flutter derivatives of the girder are extracted and the critical flutter state of the bridge is determined. The flutter mechanism is further discussed in detail based on the dynamic flow field characteristics and the work done by aerodynamic forces. Results show that the existence of central slot is in favour of the bridge flutter stability at null angle of attack. At large angles of attack, however, the flow field around the girder is quite different. Incoming flow could pass through the central slot and affect the vortex attaching to the girder. The upstream box extracts energy from the wind flow more easily than the downstream one, which may drive the bridge to torsional flutter instability at lower wind speeds.

Numerical investigation of wave forces on two side-by-side non-identical boxes in close proximity under wave actions

Wave forces on two side-by-side non-identical boxes in close proximity under wave actions are investigated by employing a numerical wave flume based on the OpenFOAM® package. The similarity and discrepancy of hydrodynamic behavior between the wave response in the narrow gap and the wave forces on the boxes are the focus of the present study. Around resonant frequencies, the large-amplitude piston-type free surface oscillation in the narrow gap can lead to the peak values in the horizontal and vertical wave forces on the downstream box, and the horizontal wave forces on the upstream box. However, only a rapid decrease with the incident wave frequencies can be observed for the vertical wave forces on the upstream box. The resonant frequencies of the wave forces on two boxes are also different with those of wave response in the narrow gap. With the increase of incident wave amplitude, the resonant frequencies and normalized amplitudes of wave forces on downstream box tend to be smaller, which is similar with that of wave response in the narrow gap. However, the normalized wave forces on the upstream box around resonant frequencies do not always decrease with the increase of incident wave amplitude. On the whole, the hydrodynamic behavior of wave forces has some similar characteristics with that of wave response in the narrow gap. However, evident discrepancy between them can also be observed because the other factors, such as the wave response upstream and downstream the two-box systems, also has the non-negligible contribution to wave forces.

Effects of gap width on flow motions around twin-box girders and vortex-induced vibrations

Twin-box girders are employed in long-span suspension bridges for flutter stability. However, the flow around a twin-box girder is notably complicated. The effects of gap width on the flow characteristics at Re=5.58×104 and the vortex-induced vibration (VIV) of a twin-box girder at about Re=1.17×104 were experimentally investigated using a sectional model. In the experiments, the gap ratio (defined as the ratio of the width L between two box girders to the central height of the girder D) varied from 0.855 to 10.260. The flow velocities around the model and the surface pressures of the model were measured during static tests, and the acceleration of sectional model also measured during dynamic tests. In addition, corresponding smoke-wire flow visualization tests were conducted under low wind speeds for the same sectional model. The test results show that all of the pressure characteristics, the aerodynamic force, the Strouhal number, the flow pattern and the VIV are dramatically influenced by the gap width. At a gap ratio of L/D=2.138 (defined as the critical gap ratio), the flow motion around the gap between the upstream and downstream box girders suddenly changes, and an alternate shedding vortex appears. According to vortex shedding pattern of the twin-box girder, the flow can be categorized into three basic patterns: pattern A, vortices alternately shedding behind the downstream box girder when the gap ratio is smaller than the critical gap ratio (L/D<2.138); pattern B, vortices alternately shedding behind the upstream box girder (in the gap) when the gap ratio is in the moderate range (2.138≤L/D≤10.26); and pattern C, vortices freely shedding behind the upstream and downstream box girders when the gap ratio approaches infinity. For the VIVs, the twin-box girder experiences four types of VIV as the gap ratio increases: when L/D<1.710, no VIV occurs; when 1.710≤L/D≤2.138, only vertical VIV occurs; when 2.566≤L/D≤2.933, both vertical and torsional VIVs occur; and when L/D≥3.421, only torsional VIV forms.

On the effects of the gap on the unsteady pressure characteristics of two-box bridge girders

This experimental study focuses on the unsteady pressure characteristics of two-box bridge decks. Models with vertical plates and gratings in several gap length configurations are investigated and compared with single-box cross sections. The results demonstrate that, with respect to unsteady pressure characteristics, the upstream box is barely affected by the slot and behaves similarly to the single-box model. In contrast, the unsteady pressure characteristics of the dowstream box are defined mainly by the characteristics of the slot and the geometry of its cross section. The upstream box geometric configuration plays only a minor role in the definition of the unsteady pressure characteristics of the dowstream box. An equivalent Theodorsen function to be used in the calculation of aerodynamic derivatives of two-box girders is proposed. Using this function, it is demonstrated that the only aerodynamic derivative that is aerodynamically dependent on the gap length is A2∗. These findings suggest that the results of single-box investigations can be extended to two-box cross sections.

Antibiotics, arsenate and H2O2 induce the promoter of Staphylococcus aureus cspC gene more strongly than cold

Proteins expressed by the bacterial cold shock genes are highly conserved at sequence level and perform various biological functions in both the cold-stressed and normal cells. To study the effects of various agents on the cold shock genes of Staphylococcus aureus, we have cloned the upstream region of cspC from S. aureus Newman and found that the above region possesses appreciable promoter (P(c)) activity even at 37 degrees C. A reporter S. aureus strain CHANDA2, constructed by inserting the P(c)-lacZ transcriptional fusion into S. aureus RN4220 genome, was found to express very low level of beta-galactosidase after cold shock, indicating that low temperature induces P(c) very weakly. Interestingly, transcription from P(c ) was induced very strongly by several antibiotics, hydrogen peroxide and arsenate salt. Cold shock proteins expressed by S. aureus are highly identical at sequence level and bear single-strand nucleic acid binding motifs. A 16 nt downstream box and a 13 nt upstream box were identified at the downstream of initiation codon and at the upstream of ribosome binding site of csp transcripts. Their roles in S. aureus cold shock gene expression have been discussed elaborately.

Mode of action of the TyrR protein: repression and activation of the tyrP promoter of Escherichia coli.

The tyrP gene of Escherichia coli encodes a tyrosine specific transporter. Its synthesis is repressed by tyrosine but is activated by phenylalanine and to a lesser extent by tryptophan. Both of these effects are mediated by the TyrR protein when it binds to one or both of its cognate binding sites (TyrR boxes) which encompass nucleotides -30 to -75. Activation in the presence of phenylalanine or tryptophan involves a dimer binding to the upstream box and interacting with the alpha subunit (alphaCTD) of RNA polymerase (RNAP). Repression in the presence of tyrosine involves a hexamer binding to both TyrR boxes. The molecular basis for this repression has been studied in vitro. Whereas initial gel shift experiments fail to show the exclusion of RNAP from the promoter region when TyrR hexamer is bound, a DNase I analysis of slices from the gel shows that in the presence of TyrR, RNAP now binds to a previously unrecognized upstream promoter. Although this upstream promoter is bound strongly by RNAP and forms an open complex on linear DNA templates, it fails to form an open complex on supercoiled templates in vitro and is unable to initiate transcription in vivo. A subsequent gel shift assay using a tyrP fragment which eliminates the upstream RNAP binding site confirms conclusively that, in the presence of tyrosine and ATP, the TyrR protein prevents RNAP from binding to the tyrP promoter. In vitro studies have also been carried out in the presence of TyrR protein and phenylalanine. Binding of TyrR protein to the upstream TyrR box in the presence of phenylalanine is shown to increase the affinity of RNAP for the promoter and stimulate open complex formation at the -10 region of the tyrP promoter. This observation coupled with the results from mutational analysis supports the proposal that TyrR-phenylalanine activates tyrP transcription by stimulating the onset of open complex formation.

Sp1 and finb interact with BETA2/NeuroD to enhance its transactivation of the secretin promoter

of full length fragment which covers all three gastrin responsive elements were cloned for yeast one hybrid screening. Electrophoretic mobility shift assays (EMSAs) were then used to confirm the positives. To determine the functional importance of the target factor(s), transient transfection and reporter assays were performed by overexpression of the target nuclear factor(s) in the presence of the HDC promoter construct which contains 125bp upstream sequence and 126bp downstream sequence followed by the luciferase gene. Results: Yeast one hybrid screening indicated that gut-enriched Kruppel like factor (GKLF) binds downstream gastrin responsive elements of the HDC promoter. Electrophoretic mobility shift assays confirmed the binding of GKLF with all three downstream gastrin responsive elements. EMSA competition assay also showed the binding of Spl with all three gastrin responsive elements. Furthermore, EMSAs indicated that GKLF binds Spl bound GC-box fragment which is located upstream of the HDC promoter. Overexpression of GKLF and Spl transactivated HDC promoter activity, which was further supported by the repressive effect o f their dominant negative plasmids. Conclusion: GKLF and Sp 1 synergistically regulate HDC promoter activity through binding both upstream GC box and downstream gastrin responsive elements.

Nonsense Mutations in cspA Cause Ribosome Trapping Leading to Complete Growth Inhibition and Cell Death at Low Temperature in Escherichia coli

CspA, the major cold shock protein of Escherichia coli, is dramatically induced immediately after cold shock. CspA production is transient and reduces to a low basal level when cells become adapted. Here we show that expression from multicopy plasmids of mutant cspA mRNAs bearing nonsense mutations in the coding region caused sustained high levels of the mutant mRNAs at low temperature, resulting in complete inhibition of cell growth ultimately leading to cell death. We demonstrate that the observed growth inhibition was caused by largely exclusive occupation of cellular ribosomes by the mutant cspA mRNAs. Such sequestration of ribosomes even occurs without a single peptide bond formation, implying that the robust translatability of the cspA mRNA is determined at the step of initiation. Further analysis demonstrated that the downstream box of the cspA mRNA was dispensable for the effect, whereas the upstream box of the mRNA was essential. Our system may offer a novel means to study sequence or structural elements involved in the translation of the cspA mRNA and may also be utilized to regulate bacterial growth at low temperature.

Localization of an exonic splicing enhancer responsible for mammalian natural trans-splicing.

Carnitine octanoyltransferase (COT) produces three different transcripts in rat through cis- and trans-splicing reactions, which may lead to the synthesis of two proteins. Generation of the three COT transcripts in rat does not depend on sex, development, fat feeding, the inclusion of the peroxisome proliferator diethylhexyl phthalate in the diet or hyperinsulinemia. In addition, trans-splicing was not detected in COT of other mammals, such as human, pig, cow and mouse, or in Cos7 cells from monkey. Rat COT exon 2 contains two purine-rich sequences. Mutation of the rat COT exon 2 upstream box does not affect the trans-splicing in vitro between two truncated constructs containing exon 2 and its adjacent intron boundaries. In contrast, mutation of the downstream box from the rat sequence (GAAGAAG) to a random sequence or the sequence observed in the other mammals (AAAAAAA) decreased trans-splicing in vitro. In contrast, mutation of the AAAAAAA box of human COT exon 2 to GAAGAAG increases trans-splicing. Heterologous reactions between COT exon 2 from rat and human do not produce trans-splicing. HeLa cells transfected with minigenes of rat COT sequences produced cis- and trans-spliced bands. Mutation of the GAAGAAG box to AAAAAAA abolished trans-splicing and decreased cis-splicing in vivo. We conclude that GAAGAAG is an exonic splicing enhancer that could induce natural trans-splicing in rat COT.

Mutation analysis of the 5' untranslated region of the cold shock cspA mRNA of Escherichia coli.

The mRNA for CspA, a major cold shock protein in Escherichia coli, contains an unusually long (159 bases) 5' untranslated region (5'-UTR), and its stability has been shown to play a major role in cold shock induction of CspA. The 5'-UTR of the cspA mRNA has a negative effect on its expression at 37 degrees C but has a positive effect upon cold shock. In this report, a series of cspA-lacZ fusions having a 26- to 32-base deletion in the 5'-UTR were constructed to examine the roles of specific regions within the 5'-UTR in cspA expression. It was found that none of the deletion mutations had significant effects on the stability of mRNA at both 37 and 15 degrees C. However, two mutations (Delta56-86 and Delta86-117) caused a substantial increase of beta-galactosidase activity at 37 degrees C, indicating that the deleted regions contain a negative cis element(s) for translation. A mutation (Delta2-27) deleting the highly conserved cold box sequence had little effect on cold shock induction of beta-galactosidase. Interestingly, three mutations (Delta28-55, Delta86-117, and Delta118-143) caused poor cold shock induction of beta-galactosidase. In particular, the Delta118-143 mutation reduced the translation efficiency of the cspA mRNA to less than 10% of that of the wild-type construct. The deleted region contains a 13-base sequence named upstream box (bases 123 to 135), which is highly conserved in cspA, cspB, cspG, and cspI, and is located 11 bases upstream of the Shine-Dalgarno (SD) sequence. The upstream box might be another cis element involved in translation efficiency of the cspA mRNA in addition to the SD sequence and the downstream box sequence. The relationship between the mRNA secondary structure and translation efficiency is discussed.