Light pollution may create demographic traps for nocturnal insects

Light pollution impacts both intra- and inter-specific interactions, such as interactions between mates and predator–prey interactions. In mobile organisms attracted to artificial lights, the effect of light pollution on these interactions may be intensified. If organisms are repelled by artificial lights, effects of light pollution on intra- and inter-specific interactions may be diminished as organisms move away. However, organisms repelled by artificial lights would likely lose suitable habitat as light pollution expands. Thus, we investigated how light pollution affects both net attraction or repulsion of organisms and effects on intra- and inter-specific interactions. In manipulative field studies using fireflies, we found that Photuris versicolor and Photinus pyralis fireflies were lured to artificial (LED) light at night and that both species were less likely to engage in courtship dialogues (bioluminescent flashing) in light-polluted field plots. Light pollution also lowered the mating success of P. pyralis. P. versicolor is known to prey upon P. pyralis by mimicking the flash patterns of P. pyralis, but we did not find an effect of light pollution on Photuris–Photinus predator–prey interactions. Our study suggests, that for some nocturnal insects, light-polluted areas may act as demographic traps, i.e., areas where immigration exceeds emigration and inhibition of courtship dialogues and mating reduces reproduction. Examining multiple factors affecting population growth in concert is needed to understand and mitigate impacts of light pollution on wildlife.     

Red- and green-emitting firefly luciferase mutants for bioluminescent reporter applications

Light emission from the North American firefly Photinus pyralis, which emits yellow-green (557-nm) light, is widely believed to be the most efficient bioluminescence system known, making this luciferase an excellent tool for monitoring gene expression. Here, we present studies leading to the production of a set of red- and green-emitting luciferase mutants with bioluminescent properties suitable for expanding the use of the P. pyralis system to dual-color reporter assays, biosensor measurements with internal controls, and imaging techniques. Using a combination of mutagenesis methods, we determined that the Ser284Thr mutation was sufficient to create an excellent red-emitting luciferase with a bioluminescence maximum of 615 nm, a narrow emission bandwidth, and favorable kinetic properties. Also, we developed a luciferase, containing the changes Val241Ile, Gly246Ala, and Phe250Ser, whose emission maximum was blue-shifted to 549 nm, providing a set of enzymes whose bioluminescence maxima were separated by 66 nm. Model studies demonstrated that in assays using a set of optical filters, the luciferases could be detected at the attomole level and seven orders of magnitude higher. In addition, in the presence of the Ser284Thr enzyme serving as a control, green light emission could be measured over a 10,000-fold range. The results presented here with the P. pyralis mutants provide evidence that simultaneous multiple analyte assay development is feasible with these novel proteins that require only a single substrate.     

An enhanced chimeric firefly luciferase-inspired enzyme for ATP detection and bioluminescence reporter and imaging applications

Firefly luciferases, which emit visible light in a highly specific ATP-dependent process, have been adapted for a variety of applications, including gene reporter assays, whole-cell biosensor measurements, and in vivo imaging. We previously reported the approximately 2-fold enhanced activity and 1.4-fold greater bioluminescence quantum yield properties of a chimeric enzyme that contains the N-domain of Photinus pyralis luciferase joined to the C-domain of Luciola italica luciferase. Subsequently, we identified 5 amino acid changes based on L. italica that are the main determinants of the improved bioluminescence properties. Further engineering to enhance thermal and pH stability produced a novel luciferase called PLG2. We present here a systematic comparison of the spectral and physical properties of the new protein with P. pyralis luciferase and demonstrate the potential of PLG2 for use in assays based on the detection of femtomole levels of ATP. In addition, we compared the performance of a mammalian codon-optimized version of the cDNA for PLG2 with the luc2 gene in HEK293T cells. Using an optimized low-cost assay system, PLG2 activity can be monitored in mammalian cell lysates and living cells with 4.4-fold and approximately 3.0-fold greater sensitivity, respectively. PLG2 could be an improved alternative to Promega’s luc2 for reporter and imaging applications.     

Characteristics of luciferases from a variety of firefly species: Evidence for the presence of luciferase isozymes

The properties of luciferases from five species of adult fireflies have been compared. The pI values as determined by isoelectric focusing are essentially the same for all five enzymes. All of the enzymes are inhibited to a comparable extent by antibody raised against the enzyme from Photinus pyralis. These experiments suggest that the enzyme found in various adult species is similar.An isozyme of luciferase, distinct from the adult form, exists in the larval stage of the firefly, Photuris pennsylvanica. Michaelis constants, pH profiles, and molecular weights are similar for larval and adult luciferases. However, the isoelectric points and the antigenicity of the larval and adult luciferases are significantly different.     

Novel Cyanobacterial Biosensor for Detection of Herbicides

The aim of this work was to generate a cyanobacterial biosensor that could be used to detect herbicides and other environmental pollutants. A representative freshwater cyanobacterium, Synechocystis sp. strain PCC6803, was chromosomally marked with the luciferase gene luc (from the firefly Photinus pyralis) to create a novel bioluminescent cyanobacterial strain. Successful expression of the luc gene during growth of Synechocystis sp. strain PCC6803 cultures was characterized by measuring optical density and bioluminescence. Bioluminescence was optimized with regard to uptake of the luciferase substrate, luciferin, and the physiology of the cyanobacterium. Bioassays demonstrated that a novel luminescent cyanobacterial biosensor has been developed which responded to a range of compounds including different herbicide types and other toxins. This biosensor is expected to provide new opportunities for the rapid screening of environmental samples or for the investigation of potential environmental damage.     

Allometry and proximate mechanisms of sexual selection in photinus fireflies, and some other beetles

Ever since Darwin identified it as the force responsible forthe evolution of exaggerated male characters, sexual selectionhas been the focus of research aimed at understanding the mostbizarre and intriguing morphologies and behaviors in Nature.Two congeneric species in the firefly genus Photinus, P. pyralisand P. macdermotti, afford a unique opportunity to examine theinteraction between sexual and countervailing natural selectionthat act to shape the evolution of mating behavior and bodysize in closely related species with very different courtshipstrategies. Photinus pyralis males emit very bright flashesduring their extended patrolling flights and form ‘loveknots’ of competing suitors, while P. macdermotti males,whose courtships seldom exceed two competing males, produceweaker flashes during a shorter patrolling period. Possibly as a consequence of their scramble competition andlong flights, not only is there an extremely wide range of bodysizes in P. pyralis males, but they also exhibit wing allometricslopes greater than one, and lantern allometries less than one.In contrast, P. macdermotti males do not have allometric slopessignificantly different from one. Small males, when tested inan artificial scramble situation move faster than large males,an advantage in the intense competition that ensues once a femaleis located. Females answer several males in alternation andfail to respond to all male flashes, with the evident consequencethat love knots frequently develop around such fickle females.Allometric relationships in two non-luminescent beetle specieswith non-visual courtship protocols are compared. Visual, sexuallyselected characters showed positive allometric slopes, whilenon-visual characters showed isometry or negative slopes. Datapresented here support the existence of distinct patterns inmodality-specific sexual selection.     

The nuclear factor κB inhibitor (E)-2-fluoro-4′-methoxystilbene inhibits firefly luciferase

Photinus pyralis (firefly) luciferase is widely used as a reporter system to monitor alterations in gene promoter and/or signalling pathway activities in vitro. The enzyme catalyses the formation of oxyluciferin from D-luciferin in an ATP-consuming reaction involving photon emission. The purpose of the present study was to characterize the luciferase-inhibiting potential of (E)-2-fluoro-4′-methoxystilbene, which is known as a potent inhibitor of the NF-κB (nuclear factor κB) signalling pathway that is used to modulate the NF-κB signalling pathway in vitro. Results show that (E)-2-fluoro-4′-methoxystilbene effectively inhibits firefly luciferase activity in cell lysates and living cells in a non-competitive manner with respect to the luciferase substrates D-luciferin and ATP. By contrast, the compound has no effect on Renilla and Gaussia luciferases. The mechanism of firefly luciferase inhibition by (E)-2-fluoro-4′-methoxystilbene, as well as its potency is comparable to its structure analogue resveratrol. The in vitro use of trans-stilbenes such as (E)-2-fluoro-4′-methoxystilbene or resveratrol compromises firefly luciferase reporter assays as well as ATP/luciferase-based cell viability assays.     

Roles of trehalose and magnesium sulfate on structural and functional stability of firefly luciferase

Firefly luciferase is widely used in many analytical techniques. However, the enzyme is unstable, so that its relative inactivation results in low sensitivity of those techniques. In this study, we have investigated the effects of MgSO₄ and trehalose on the structural stability and function of luciferase from Photinus pyralis using circular dichroism (CD), conventional and stopped-flow fluorescence spectroscopy and bioluminescence assay. The secondary structural content, compactness and its melting temperature are also studied, which showed that the stability of luciferase increased in the presence of additives. Measurements of refolding rate constants under conditions that favor folding, show that MgSO₄ accelerates the folding of enzyme, on the contrary, refolding rate constant decreases in the presence of trehalose which can be attributed to its high viscosity. Finally, combined with remaining activity assay we concluded that magnesium sulfate and trehalose can be used for short- and long-term stabilization, respectively.     

Bioluminescent Aspergillus fumigatus, a New Tool for Drug Efficiency Testing and In Vivo Monitoring of Invasive Aspergillosis

Aspergillus fumigatus is the main cause of invasive aspergillosis in immunocompromised patients, and only a limited number of drugs for treatment are available. A screening method for new antifungal compounds is urgently required, preferably an approach suitable for in vitro and in vivo studies. Bioluminescence imaging is a powerful tool to study the temporal and spatial resolutions of the infection and the effectiveness of antifungal drugs. Here, we describe the construction of a bioluminescent A. fumigatus strain by fusing the promoter of the glyceraldehyde-3-phosphate dehydrogenase gene from A. fumigatus with the luciferase gene from Photinus pyralis to control the expression of the bioluminescent reporter. A. fumigatus transformed with this construct revealed high bioluminescence under all tested growth conditions. Furthermore, light emission correlated with the number of conidia used for inoculation and with the biomass formed after different incubation times. The bioluminescent strains were suitable to study the effectiveness of antifungals in vitro by several independent methods, including the determination of light emission with a microplate reader and the direct visualization of light emission with an IVIS 100 system. Moreover, when glucocorticoid-treated immunosuppressed mice were infected with a bioluminescent strain, light emission was detected from infected lungs, allowing the visualization of the progression of invasive aspergillosis. Therefore, this new bioluminescence tool is suitable to study the in vitro effectiveness of drugs and the disease development, localization, and burden of fungi within tissues and may also provide a powerful tool to study the effectiveness of antifungals in vivo.     

Assessing Condition-dependence of Male Flash Signals in <Emphasis Type="Italic">Photinus</Emphasis> Fireflies

Females often show a preference for exaggerated male sexual traits or courtship behaviors. Such preferences can benefit females if trait expression is correlated with male genetic quality or phenotypic condition. Previous studies of several Photinus fireflies have revealed considerable intraspecific variation in the bioluminescent courtship signals emitted by males, and have also demonstrated that females prefer more conspicuous male signals. Thus, females might gain information about male phenotypic quality if courtship signals reflect male condition. We examined possible condition-dependence of Photinus male courtship signals using two complementary approaches. First we experimentally manipulated male mating status, which is expected to affect male condition by depleting resources required for nuptial gift formation, and looked at how individual male flash signals changed with mating status and over time. We used an additional approach to assess condition-dependence by examining whether a relationship exists between flash signal parameters and measures of male condition and body shape. We found that the pulse rate of P. greeni courtship signals was not altered by male mating status or age, and that the pulse duration of P. ignitus signals was also not affected by male mating status. In P. pyralis fireflies, males showed a non-significant trend toward reduced signal pulse duration with age. When we examined the relationship between male flash signals and condition measures, we found no effect of male condition or body shape on courtship signals in P. greeni or P. ignitus; in P. pyralis, males with wider body shapes produced longer duration flash signals. On the other hand, we found no evidence in P. pyralis that condition predicted flash duration. Taken together, these results indicate that Photinus males’ flash signals do not reflect adult male condition, and suggest that females are unlikely to use courtship signals as an indicator of male phenotypic quality.     

Etiology of Childhood Infectious Diarrhea in a Developed Region of China: Compared to Childhood Diarrhea in a Developing Region and Adult Diarrhea in a Developed Region

In China, great differences in economy, social characteristics and hygiene exist between developing and developed regions. A comparative study of infectious diarrhea between two regions was needed. Three groups of diarrheal patients were collected: children ≤5 year-olds from Beijing (developed region) and Henan Province (developing region), and adults over 18 year-olds from Beijing. A questionnaire was used to survey and feces samples were examined for 16 enteropathogens. We enrolled 1422 children and 1047 adults from developed region and 755 children from developing region. Virus positive rates were 32.98% for children and 23.67% for adults in developed region. The most prevalent pathogen for children was rotavirus whereas for adults was norovirus. Bacterial isolation rates were 13.92% for children from developed region, while 29.14% for children from the developing regions. For the greatest difference, Shigella accounted for 50.79% and was the dominant pathogen in the developing region, whereas in the developed region it was only 1.45%. There was no significant relationship between the local levels of development with diarrheogenic Escherichia coli (DEC) categories. But it was seen the notable differences between the population with different age: enteropathogenic E.coli (EPEC) and enteroaggregative E.coli (EAggEC) were the primary classes of DEC in children from both regions, whereas it was enterotoxigenic E.coli (ETEC) in adults. The symptoms of Shigella and Salmonella infection, such as bloody stools, white blood cells (WBC) and red blood cells (RBC) positivity and fever were similar in children, which may lead to the misidentification. Yersinia enterocolitica and shiga toxin-producing E.coli (STEC) infections were firstly reported in Beijing. There was a large difference in etiology of bacterial diarrhea between children in developing and developed regions of China.     

Aberrant 5’-CpG Methylation of Cord Blood TNFα Associated with Maternal Exposure to Polybrominated Diphenyl Ethers

Growing evidence suggests that maternal exposures to endocrine disrupting chemicals during pregnancy may lead to poor pregnancy outcomes and increased fetal susceptibility to adult diseases. Polybrominated diphenyl ethers (PBDEs), which are ubiquitously used flame-retardants, could leach into the environment; and become persistent organic pollutants via bioaccumulation. In the United States, blood PBDE levels in adults range from 30–100 ng/g- lipid but the alarming health concern revolves around children who have reported blood PBDE levels 3 to 9-fold higher than adults. PBDEs disrupt endocrine, immune, reproductive and nervous systems. However, the mechanism underlying its adverse health effect is not fully understood. Epigenetics is a possible biological mechanism underlying maternal exposure-child health outcomes by regulating gene expression without changes in the DNA sequence. We sought to examine the relationship between maternal exposure to environmental PBDEs and promoter methylation of a proinflammatory gene, tumor necrosis factor alpha (TNFα). We measured the maternal blood PBDE levels and cord blood TNFα promoter methylation levels on 46 paired samples of maternal and cord blood from the Boston Birth Cohort (BBC). We showed that decreased cord blood TNFα methylation associated with high maternal PBDE47 exposure. CpG site-specific methylation showed significantly hypomethylation in the girl whose mother has a high blood PBDE47 level. Consistently, decreased TNFα methylation associated with an increase in TNFα protein level in cord blood. In conclusion, our finding provided evidence that in utero exposure to PBDEs may epigenetically reprogram the offspring’s immunological response through promoter methylation of a proinflammatory gene.     

Downstream processing of luciferase from fireflies (Photinus pyralis) using aqueous two-phase extraction

The firefly luciferase has been extensively used for sensitive detection of bacteria, gene expression and environmental toxins (biosensors). The aim of the present study was to design a simple and more efficient method for the purification and concentration of luciferase using aqueous two-phase extraction (ATPE). Downstream processing of luciferase from North American Firefly Photinus pyralis was carried out, for the first time, using polymer/salt aqueous two phase system (ATPS) at 4 °C. The enzyme was observed to preferentially partition to the polyethylene glycol (PEG) rich top phase. The best results of purification (13.69 fold) and enzyme activity recovery (118.34%) were observed in the system containing 4.0% (w/w) PEG (1500) and 20.5% (w/w) (NH₄)₂SO₄ with a phase volume ratio of 0.21.     

Bartonella Infection among Cats Adopted from a San Francisco Shelter,Revisited

Bartonella infection among cats from shelters can pose a health risk to adopters. Bartonella henselae is the most common species, with B. clarridgeiae and B. koehlerae being less common. The lower rates of infection by the latter species may reflect their rarity or an inefficiency of culture techniques. To assess the incidence of infection, blood cultures, serology, and PCR testing were performed on 193 kittens (6 to 17 weeks old) and 158 young adult cats (5 to 12 months old) from a modern regional shelter. Classical B. henselae culture medium was compared to a medium supplemented with insect cell growth factors. Bartonella colonies were isolated from 115 (32.8%) animals, including 50 (25.9%) kittens and 65 (41.1%) young adults. Therefore, young adults were twice as likely to be culture positive as kittens. Enhanced culture methods did not improve either the isolation rate or species profile. B. henselae was isolated from 40 kittens and 55 young adults, while B. clarridgeiae was cultured from 10 animals in each group. B. koehlerae was detected in one young adult by PCR only. B. henselae genotype II was more commonly isolated from young adults, and genotype I was more frequently isolated from kittens. Kittens were 4.7 times more likely to have a very high bacterial load than young adults. A significantly higher incidence of bacteremia in the fall and winter than in the spring and summer was observed. Bartonella antibodies were detected in 10% (19/193) of kittens and 46.2% (73/158) of young adults, with culture-positive kittens being 9.4 times more likely to be seronegative than young adults.     

Evidence of Experimental Vertical Transmission of Emerging Novel ECSA Genotype of Chikungunya Virus in Aedes aegypti

Background

Chikungunya virus (CHIKV) has emerged as one of the most important arboviruses of public health significance in the past decade. The virus is mainly maintained through human-mosquito-human cycle. Other routes of transmission and the mechanism of maintenance of the virus in nature are not clearly known. Vertical transmission may be a mechanism of sustaining the virus during inter-epidemic periods. Laboratory experiments were conducted to determine whether Aedes aegypti, a principal vector, is capable of vertically transmitting CHIKV or not.

Methodology/Principal Findings

Female Ae. aegypti were orally infected with a novel ECSA genotype of CHIKV in the 2^nd gonotrophic cycle. On day 10 post infection, a non-infectious blood meal was provided to obtain another cycle of eggs. Larvae and adults developed from the eggs obtained following both infectious and non-infectious blood meal were tested for the presence of CHIKV specific RNA through real time RT-PCR. The results revealed that the larvae and adults developed from eggs derived from the infectious blood meal (2^nd gonotrophic cycle) were negative for CHIKV RNA. However, the larvae and adults developed after subsequent non-infectious blood meal (3^rd gonotrophic cycle) were positive with minimum filial infection rates of 28.2 (1∶35.5) and 20.2 (1∶49.5) respectively.

Conclusion/Significance

This study is the first to confirm experimental vertical transmission of emerging novel ECSA genotype of CHIKV in Ae. aegypti from India, indicating the possibilities of occurrence of this phenomenon in nature. This evidence may have important consequence for survival of CHIKV during adverse climatic conditions and inter-epidemic periods.     

A topical metabolic inhibitor to improve transcutaneous estimation of arterial oxygen tension in adults

The use of Eugenol as a contact medium in the measurement of transcutaneous oxygen (tcPO₂) partially inhibits the metabolism in the underlying tissue, thereby reducing oxygen consumption and increasing tcPO₂. Oxygen consumption in the tissue can be estimated from the rate at which tcPO₂ falls when blood flow is occluded, and blood flow in the tissue can be estimated from the rate at which tcPO₂ increases when the subject changes from breathing air to pure oxygen. Both these measurements have been made with Eugenol and distilled water as contact media. From these measurements it has proved possible to estimate the arterial oxygen tension (aPO₂) of healthy adults at a relatively low sensor temperature (43°C).     

Etiología de la candidiasis en pediatría: análisis comparativo con el paciente adulto

BackgroundCandida spp. represents a group of commensal yeasts that can act as pathogens and cause candidiasis in different anatomical locations.AimsThe aim of this study was to perform an epidemiological and comparative analysis between the isolates of Candida spp. in clinical specimens during a three year-period (2010-2012) from children (0-14 years) and adults (15-99 years) in the Valencian Community (RedMIVA).MethodsThe microbiological surveillance network of Valencian Community was used as the information source.Results and conclusionsCandida was isolated in 52,436 patients (1,604 [3.1%] children and 50,832 [96.9%] adults). Candida albicans was significantly (p < 0.05) the predominant species in both age groups, and in almost every type of clinical specimen. The distribution of other species varied depending on the sample type and age group. In blood specimens, Candida parapsilosis followed by C. albicans, Candida famata and Candida lusitaniae were the main species found in children, whereas C. albicans followed by C. parapsilosis, Candida glabrata and Candida tropicalis were the predominant species in adults. In sterile fluids, urine and lower respiratory tract samples, C. parapsilosis was the second most prevalent species in the children group, while C. glabrata and C. tropicalis were the main second species in adults.     

Whole blood interferon-gamma responses to mycobacterium tuberculosis antigens in young household contacts of persons with tuberculosis in Uganda

Background

Due to immunologic immaturity, IFN-γ-producing T cell responses may be decreased in young children compared to adults, thus we hypothesized that IFN-γ responses to mycobacterial antigens in household contacts exposed to Mycobacterium tuberculosis (Mtb) would be impaired in young children relative to adults. The objective of this study was to compare whole blood IFN-γ production in response to mycobacterial antigens between children and adults in Uganda.

Methodology/Principal Findings

We studied household contacts of persons with culture-positive pulmonary tuberculosis (TB) enrolled in a cohort study conducted in Kampala, Uganda. Whole blood IFN-γ production in response to Mtb culture-filtrate antigens was measured by ELISA and compared between infants (<2 years old, n = 80), young children (2 <5 years old, n = 216), older children (5 <15 years old, n = 443) and adults (≥15 years old, n = 528). We evaluated the relationship between IFN-γ responses and the tuberculin skin test (TST), and between IFN-γ responses and epidemiologic factors that reflect exposure to Mtb, and the effect of prior BCG vaccination on IFN-γ responses. Young household contacts demonstrated robust IFN-γ responses comparable to those of adults that were associated with TST and known risk factors for infection. There was no effect of prior BCG immunization on the IFN-γ response.

Conclusions/Significance

Young children in a TB endemic setting can mount robust IFN-γ responses generally comparable to those of adults, and as in adults, these responses correlated with the TST and known epidemiologic risk factors for Mtb infection.     

Feeding ethogram of Antilochus coquebertii F. preying on Dysdercus koenigii

Antilochus coquebertii F. (Hemiptera: Pyrrhocoridae) is one of the most important and voracious feeders of Dysdercus spp. First time, this study explained the feeding ethogram of both male and female A. coquebertii on Dysdercus koenigii adults (both male and female). Female adults of A. coquebertii consumed maximum prey (97.78 %) with respect to male (90.91%). The pre-feeding time was recorded maximum in male adults (12.22 min) as compared to female adults (4.1 min). In case of paralysis time, female adults take more time (0.47 min) as compared to male adults (0.46 min). Both male and female adults of A. coquebertii exhibited different preferences towards different body parts of D. koenigii with variable feeding time and number of feedings. Feeding ethogram showed that both of male and female A. coquebertii adults use their antennae in olfactory and gustatory sense. Based on feeding time, abdomen was more favored by male adults and thorax by female adults. While, in case of number of feedings, male adults of A. coquebertii preferred to feed on head and female on thorax of D. koenigii. Increase in body weight of female adults of A. coquebertii after feeding on its prey was found maximum (0.92 g) as compared to male adult (0.66 g). According to these findings, antennae of A. coquebertii have olfactory and gustatory sense while proboscis kept gustatory sense. Female adults of A. coquebertii is more efficient feeder of D. koenigii than male. Abdomen of D. koenigii was more preferred by male adults, while thorax by female adults of A. coquebertii.     

Characterization and Development of Two Reporter Gene Systems for Clostridium acetobutylicum

The use of lacZ from Thermoanaerobacterium thermosulfurigenes (encoding β-galactosidase) and lucB from Photinus pyralis (encoding luciferase) as reporter genes in Clostridium acetobutylicum was analyzed with promoters of genes required for solventogenesis and acidogenesis. Both systems proved to be well suited and allowed the detection of differences in promoter strength at least up to 100-fold. The luciferase assay could be performed much faster and comes close to online measurement. Resequencing of lacZ revealed a sequence error in the original database entry, which resulted in β-galactosidase with an additional 31 amino acids. Cutting off part of the gene encoding this C terminus resulted in decreased enzyme activity. The lacZ reporter data showed that bdhA (encoding butanol dehydrogenase A) is expressed during the early growth phase, followed by sol (encoding butyraldehyde/butanol dehydrogenase E and coenzyme A transferase) and bdhB (encoding butanol dehydrogenase B) expression. adc (encoding acetoacetate decarboxylase) was also induced early. There is about a 100-fold difference in expression between adc and bdhB (higher) and bdhA and the sol operon (lower). The lucB reporter activity could be increased 10-fold by the addition of ATP to the assay. Washing of the cells proved to be important in order to prevent a red shift of bioluminescence in an acidic environment (for reliable data). lucB reporter measurements confirmed the expression pattern of the sol and ptb-buk (encoding phosphotransbutyrylase and butyrate kinase) operons as determined by the lacZ reporter and showed that the expression level from the ptb promoter is 59-fold higher than that from the sol operon promoter.