Search

The low success rates in the treatment of multidrug-resistant tuberculosis (MDR-TB) and extensively drug-resistant TB (XDR-TB), which account for 55% and 34% respectively, led the WHO to conclude that MDR/XDR-TB is a serious public health crisis. However, the virulence of MDR/XDR-Mycobacterium Tuberculosis(Mtb) has not been analyzed in details, which could provide a specific guidance for the control and prevention. In this review, we discuss different aspects of MDR/XDR-Mtb virulence and its relationship to fitness cost by probing the following questions: (1) what mediates the virulence of MDR/XDR-Mtb? (What is the relationship between fitness and virulence of Mtb? (2) Is it possible that drug-resistant Mtb(DR Mtb) can show higher fitness? (3) What is the definite effect on fitness of each drug-resistant mutant? (4) What other important factors affecting fitness in the mutant strain? (5) How to study the virulence of a large number of DR Mtb?And what prevention and control measures will be taken in the future, especially for the high virulent DR Mtb? We therefore summarized the congruent relationship between drug resistance and fitness from the global response-related genes to antibiotic resistance-contributing mutation, provided methods to explore the virulence of DR Mtb. This review may offer some critical information and concise guide to creating strategies for the prevention and control of drug-resistant Mtb.

This study reports on the use of Fourier transform infrared (FTIR) microspectroscopy imaging to investigate biochemical changes occurring during C. elegans lifespan. C. elegans wild-type (N2) and the tub-1 mutant strain were cultured in agar plates. FTIR imaging was performed on single worms in transmission mode at day 8, 11, and 15. Principal component analysis was then performed to analyze the spectra acquired during C. elegans lifespan. The FTIR spectra were clustered in three groups corresponding to the spectra acquired from the worms at day 8, 11, and 15. The results showed major changes in lipids (vibration 1744 cm−1 assigned to C O stretching) and proteins (vibrations 1648 cm−1 assigned to C O stretching of amide I and 1548 cm−1 assigned to N H bending and C–N stretching in amid II). The vibration assigned to glycogen around 1155 cm−1 was present in the spectra acquired at day 8 and 11 but the peak area decreased by 49.3% and 73.3% in the spectra acquired at day 15 respectively from WT(N2) and tub-1 mutant strain. Furthermore, PC-1 loadings as a function of wavenumbers show that the presence of the vibration 1698 cm−1, attributed to antiparallel β-sheet, could indicate the formation of lipofuscin. The results obtained demonstrate that FTIR imaging could be used as a tool to monitor biochemical changes during lifespan studies, which could bring additional information to our understanding of longevity.

New indolylarylsulfone HIV-1 NNRTIs were synthesized to evaluate unexplored substitutions of the benzyl/phenylethyl group linked at the indole-2-carboxamide. Against the NL4-3 HIV-1 WT strain, 17 out 20 compounds were superior to NVP and EFV. Several compounds inhibited the K103N HIV-1 mutant strain at nanomolar concentration and were superior to EFV. Some derivatives were superior to EFV against the Y181C and L100I HIV-1 mutant strains. Against the NL4-3 HIV-1 strain, the enantiomers 24 and 25 showed small differences of activity. In contrast, 24 turned out significantly more potent than 25 against the whole panel of mutant HIV-1 strains. The docking studies suggested that the difference in the observed inhibitory activities of 24 and 25 against the K03N mutation could be due to a kinetic rather than affinity differences.

Photoelectrocatalytic (PEC) and photocatalytic (PC) inactivation of ancestor Escherichia coli K-12 and its mutant E. coli BW25113 were systematically compared using a TiO2 nanotubular photoanode. The results showed that PEC inactivation was more effective to both bacterial strains than PC process, and E. coli BW25113 showed higher resistance than E. coli K-12 in both PEC and PC systems. The findings indicate that the two strains with different genes are varied in their susceptibilities and responses to the PEC and PC treatments. The h+ was found to be the major reactive species and predominantly responsible for PEC inactivation. Scanning electron microscopy images demonstrated that the cells were severely damaged and resulted in a leakage of the intracellular components during PEC inactivation process. For a given bacterial strain, no significant effect was found on the PEC inactivation efficiency as different electrolytes were employed. However, in the presence of NaCl or NaBr, PEC inactivation efficiencies of both strains were remarkably enhanced. This phenomenon can be attributed to the efficient formation of halide and dihalide radical anions during PEC process. The different efficiencies of the two bacterial strains under same conditions can be ascribed to their different abilities to resist the inactivation of bacterial strains with different genotypes.

A series of novel N-arylmethyl substituted piperidine-linked aniline derivatives were designed, synthesized and evaluated for their anti-HIV activity in MT-4 cells. All the new compounds showed moderate to potent activities against wild-type (wt) HIV-1 with an EC50 range from 0.022 to 2.1 μM. Among them, compound 5a6 (EC50 = 0.022 ± 0.0091 μM, SI >10,770) was confirmed to be the most potent and selective inhibitor, which proved more potent than DDI and DLV in a cell-based assay against wt HIV-1, and more efficient than NVP in an RT (reverse transcriptase) assay. Besides, it is worth noting that compound 7a1 retained moderate inhibitory activity (EC50 = 4.8 ± 0.95 μM) against the HIV-1 double RT mutant strain (K103N/Y181C). The preliminary structure–activity relationship was discussed and rationalized by molecular simulation.

Natural herbicides, or environment-friendly bioherbicides have been attracted more and more attentions. Isolation and structural identification of natural herbicide-active compounds from plant pathogens has been proved to be an effective approach for novel lead discovery of the pesticide development. In this study, the metabolites of the mutant strain PAM1, which obtained from PA1 of Pythium aphanidermatum (Eds.) Fitzp by ultraviolet radiation were separated and identified by HPLC, NMR, and IR. The results revealed that three active compounds including 4-hydroxy-3-methoxycinnamic acid and two indole derivatives, exhibited inhibition activity on the elongation of radical and coleoptile of Digtaria sanguinalis (L.) Scop.

Dynamic modeling of bioprocesses is of fundamental importance for optimization and control. The underlying macroscopic models, i.e., reaction scheme stoichiometry and kinetics, have to be derived and identified from experimental data. This study addresses the problem of determining the kinetics, and proposes a systematic procedure based on the use of decision graphs collecting well-established kinetic laws, and organizing them according to their complexity (i.e., number of unknown parameters). An algorithm for selecting the most likely kinetic structures, starting from the simplest models in the decision graphs, is developed based on the concepts of nested models and likelihood ratio tests. The potential of the procedure is illustrated with the determination of the synthesis rate of a biopolymer from the mutant strain Azotobacter vinelandii utilizing glucose.

Cuticle tanning in insects involves simultaneous cuticular pigmentation and hardening or sclerotization. The dynamic mechanical properties of the highly modified and cuticle-rich forewings (elytra) from Tribolium castaneum (red flour beetle) wild-type and body color mutant strains were investigated to relate body coloration and elytral mechanical properties. There was no statistically significant variation in the storage modulus E' among the elytra from jet, cola, sooty and black mutants or between the mutants and the wild-type GA-1 strain: E' averaged 5.1 ± 0.6 GPa regardless of body color. E' is a power law function of oscillation frequency for all types. The power law exponent, n, averaged 0.032 ± 0.001 for elytra from all genotypes except black; this small value indicated that the elytra are cross-linked. Black elytra, however, displayed a significantly larger n of 0.047 ± 0.004 and an increased loss tangent (tan δ), suggesting that metabolic differences in the black mutant strain result in elytra that are less cross-linked and more pigmented than the other types. These results are consistent with the hypothesis that black elytra have a β-alanine-deficient and dopamine-abundant metabolism, leading to greater melanin (black pigment) production, probably at the expense of cross-linking of cuticular proteins mediated by N-β-alanyldopamine quinone.

Biotechnological processes for the production of (high-volume) chemicals are often based on one-stage or two-stage processes (OSPs and TSPs, respectively). Many classical bioprocesses are OSPs in which growth and product synthesis are coupled, however, OSPs often exhibit low volumetric productivities due to inherent trade-offs between biomass and product yields. Here, TSPs, where growth and production are separated, may help to improve the productivity of bioprocesses and with rapidly growing genetic toolboxes for dynamic metabolic control TSPs are becoming more attractive. However, there are only few theoretical and experimental studies systematically comparing OSPs and TSPs in terms of their process metrics (productivity, yield etc.). We have constructed E. coli strains for heterologous production of itaconate for both OSP and TSP. We used computer-aided design to build a mutant strain (ita23) for growth-coupled synthesis of itaconate. In an OSP, this strain reached the highest product yield and titer that have ever been reported for heterologous itaconate production. For increasing the volumetric productivity via a TSP, we constructed a new strain (ita36A) allowing a temperature-dependent switch from fast growth to production. Indeed, a TSP with ita36A increased the peak productivity of ita23 by 91% and the titer by 46%. Despite of this successful implementation of a TSP, we observed that the productivity, after reaching a peak value, decreased quickly in the production phase due to low substrate uptake and product synthesis rates. We therefore used mathematical models to systematically compare productivities of TSPs and OSPs. Our simulations show that the overall volumetric productivity of a TSP can in fact be lower than of a corresponding OSP due to the sharp decrease of the specific substrate uptake rate in (non-growth) TSP production phases. This raises a (often neglected) big challenge for designing competitive TSPs. We propose approaches to improve substrate utilization rates in the production phase by which TSPs really outperform OSPs. In conclusion, a careful assessment of the trade-offs between substrate uptake rates, yields, and productivity is necessary when deciding for OSP vs. TSP.

We have developed a mutant strain derived from Agrobacterium sp. ATCC 31750, which produces a water-soluble polysaccharide having potential utility to the food, feed, pharmaceutical and cosmetic industries. A high concentration of product (15 g/L) is obtained by 48 h cultivation of the mutant strain under optimized fermentation conditions. The water-soluble polysaccharide obtained from cultures of the mutant strain beta82 has Glc:Man:Gal in approximate molar ratios of 5.8:6.7:1.0. The molecular weight of the polysaccharide was determined to be approximately 1000 kDa by HPSEC analysis. Linkage analysis contained 3-Glc p , 3-Man p , terminal Glc p and terminal Man p , as well as a small proportion of 3- and 3,4-Gal p , and 4,6-Man p residues. Based on analyses using FT-IR and 13 C NMR spectrometers, most glycosidic bonds joining these sugar residues are of the α-type, and acetyl groups are apparently attached to the polymer chain at random.

The biodegradations of phenol and 4-chlorophenol (4-cp) were studied using the mutant strain CTM 2 obtained by the He-Ne laser irradiation on wild-type Candida tropicalis. The results showed that the capacity of the CTM 2 to biodegrade 4-cp was increased up to 400 mg·L−1 within 59.5 h. In the dual-substrate biodegradation, both velocity and capacity of the CTM 2 to degrade 4-cp increased with low-concentration phenol. A total of 400 mg·L−1 4-cp was completely degraded within 50.5 h in the presence of 300 mg·L−1 phenol. The maximum 4-cp biodegradation could reach 440 mg·L−1 with 120 mg·L−1 phenol. Low-concentration 4-cp caused great inhibition on the CTM 2 to degrade phenol. In addition, the kinetic behaviors were described using the kinetic model proposed in this lab.

Taxa included in the Aspergillus niger aggregate are difficult to distinguish by phenotypic characterization. In this work, the effect of gentian violet on the growth of the N and T RFLP types of A. niger aggregate strains has been investigated. In total, 105 strains from different sources and origins, including reference cultures and field isolates were studied. Type N and T RFLP patterns, ochratoxin A production and the effect of different concentrations of gentian violet on the growth were determined in these strains. Forty nine strains belonged to the N type and 56 strains to the T type. Sixteen out of the 105 strains assayed were OTA producers. All the OTA-producing species belonged to the RFLP type N and none of the T type strains was able to produce OTA. Approximately 90% of the N type strains grew in the presence of 25 ppm of gentian violet. Only five N type strains did not grow on this medium. One of these strains was A. niger ATCC 22343, a well documented induced mutant strain and the remaining four strains belonged to the new species A. brasiliensis. On the contrary, all the T type strains failed to grow on this medium after 3 days of incubation (sensitivity 89.79%; specificity 100%). The use of growth in gentian violet as an additional character for classification and identification purposes in this taxonomic group may be useful because no phenotypic methods have yet been found that can distinguish between these species.

Genetic networks are often affected by stochastic noise, due to the low number of molecules taking part in certain reactions. In complex regulatory networks, noise in any one chemical species may induce noise in the rest of the system. In this paper, we analyse the sources of stochastic noise in the yeast galactose utilization pathway at the level of the complete system, by using both computer simulations, and experimental comparisons between wild-type yeast and a modified strain. A computer model was first used to determine the sources of network noise, as well as mechanisms by which noise is controlled. Results from an estimation tool, confirmed by detailed stochastic simulations, show that noise is caused primarily by fluctuations in mRNA concentrations due to their stochastic creation and decay. The noise is controlled by feedback loops which regulate transcription of certain genes. To test the effect of the feedback loops on the rest of the system, a modified strain of yeast was prepared in which regulation of two key genes is eliminated. As predicted by the model, the mutant strain is induced by galactose in a manner similar to that for the wild-type, but with a higher degree of stochastic noise.

Stipe elongation in fruit bodies of Coprinopsis cinerea (syn. Coprinus cinereus) was examined from a biomechanical perspective. Two strains were studied: the self-compatible Amut Bmut homokaryon that produces normal fruit bodies with relatively short stipes, and mutant B1918 that produces abnormally elongated stipes. Measurements of the pressure exerted by developing mushrooms were made using strain gauges, and these data were compared with measurements of the pressures exerted by vegetative hyphae of the same strains. The experiments demonstrate that AmutBmut hyphae elongating within stipe tissue push with the same pressure (approx. 0.5 atmosphere) as vegetative hyphae growing through their food sources. In purely biomechanical terms, the fruit body may therefore be viewed as a relatively uncomplicated sum of its parts. Analysis of the mutant strain B1918 demonstrated that hyperelongation of the stipe is not associated with any difference in the pressure exerted by the fruit body. The fault in the mechanism of stipe extension in B1918 may be reflected in the increased fluidity of the cell wall of vegetative hyphae of this strain, but further work is necessary to resolve this.

Beef-processing equipment can be contaminated with pathogens such as Escherichia coli O157:H7 and Salmonella spp. The bowl cutter has wide application in particle-size reduction and blending of meat products. This study was undertaken to determine (i) the distribution patterns of E. coli O157:H7 in equipment components and ground beef produced with a table-top bowl cutter under different operational conditions and (ii) the likelihood that pathogen contamination can be transferred to subsequent batches after a batch of beef contaminated with E. coli O157:H7 has been processed in the same bowl cutter. A beef trim (44.6 +/- 29.5 g) inoculated with 2 log CFU of an E. coli O157:H7 mutant strain resistant to rifampicin (E. coli O157:H7rif) was fed by hand into an uncontaminated beef-trim batch under two different batch sizes (2 and 4 kg), three processing times (60, 120, and 240 s), and two feeding modes (running and stoppage fed). There were no significant differences (P > or = 0.05) among all the treatments for the averages of the counts of E. coli O157:H7rif distributed in the ground beef. Regardless of the processing time and the method used to feed the beef trims into the bowl cutter, the whole batch and the following subsequent batch became contaminated when previously contaminated beef was processed. Areas of the bowl cutter most likely to be contaminated with E. coli O157:H7 were (i) the material left on the top of the comb/knife guard and (ii) the knife. Material that overflowed the bowl cutter, when processing the batch with E. coli O157:H7rif, contaminated the equipment surroundings. A Pearson V probability distribution function was determined to describe the distribution of pathogenic organisms in the ground beef, a distribution that can also be applied when conducting process risk analyses on mixing-particle reduction operations for beef trims.

Production of neomycin by a mutant strain of Streptomyces marinensis, under solid state fermentation was optimised. Various substrates were used to study their effect on neomycin production. Wheat rawa was found to be better for optimum production of neomycin. The maximum neomycin yield (10 755 μg/g substrate) was achieved with optimised process parameters such as coarse size of wheat rawa with raspberry seed powder (10% w/w) as additional nutrient, initial moisture content of solid substrate 80%, initial pH of the medium 7.5, incubation temperature 30 °C, inoculum size 0.5% w/w dry cell mass equivalent and 10 days of fermentation time. One hundred and forty percent of higher production was achieved after optimising the process parameters.

The recent characterization of a mutant strain of mice generated five decades ago in a program to study ionizing radiation in mammals (and, ironically, derived from a nonmutagenized control animal) is helping to dissect a now resurgent area of immunology. Despite a vast literature during the 1970s, the study of suppressor T cells had been largely abandoned until the publication of several seminal papers rekindled interest in what are today generally referred to as regulatory T (TR) cells. The identification of the transcription factor Foxp3 as the gene responsible for the defect in scurfy mice, and subsequently, the demonstration of its critical involvement in the generation of TR cells, provides an important molecular insight into this essential cell lineage.

Formylation of the initiator methionyl-tRNA, catalyzed by methionyl-tRNA formyltransferase, has long been regarded as essential for initiation of protein synthesis in eubacteria. Here, we show that this process is, in fact, dispensable in Pseudomonas aeruginosa. Disruption of the chromosomal methionyl-tRNA formyltransferase gene in P. aeruginosa resulted only in a moderate decrease in the rate of cell growth, whereas in Escherichia coli cell growth was severely impaired. The ability of the P. aeruginosa mutant strain to grow was not due to an additional copy of the methionyl-tRNA formyltransferase gene or to N-acylation of the methionyl moiety by a group other than formyl. These results indicate that P. aeruginosa can carry out formylation-independent initiation of protein synthesis, using the nonformylated methionyl-tRNA. Therefore, the dogma that eubacteria require formylation of the initiator methionyl-tRNA for initiation of protein synthesis may have been an invalid generalization of results obtained with E. coli.

A mutant strain of Salmonella typhimurium, SJW46, has flagellar filaments supercoiled in the same form as the wild-type strain, SJW1103, and swims normally. However, its flagellar filaments are mechanically unstable and show anomalous behaviors of polymorphism. Flagellin from SJW46 has a large central deletion from Ala204 to Lys292 of SJW1103 flagellin, which has been thought to be located in the outer surface of the filament. Since the filament structure is determined by intersubunit interactions of the terminal regions in the densely packed core of the filament, no serious involvement of the deleted portion was expected in the filament stability and polymorphism. In order to locate the deleted portion and to understand the underlying mechanism of these anomalous characteristics, we carried out structure analysis of the L-type straight filament reconstituted from a mutant flagellin of SJW46 (SJW46S) and compared the structure with that of the SJW1660 filament, which is also the L-type but composed of flagellin with no deletion. The deleted portion was identified as the outermost subdomain, and the structure in the core region showed no appreciable differences. The structure revealed the previously identified folding of flagellin in further detail, and the significance of intersubunit interactions between outer domains, which are present in the SJW1660 filament but absent in the SJW46 filament. This suggests that these contacts have a significant contribution to the filament stability and polymorphic behavior, despite the fact that the contacting surface area occupies only a minor portion of the whole intersubunit interactions.

Rice plants were grown in a temperature-controlled greenhouse in 1.2 L glass fleakers. The rooting media were inoculated with either the parent strain or a nitrogenase-derepressed mutant strain (which excreted NH4 4 + produced by nitrogenase) of the cyanobacterium Anabaena variabilis and exposed to 15 N 2 . Dry matter and total N accumulated in the roots and shoots of plants inoculated with the mutant strain were significantly greater than from plants inoculated with the parent strain. Significantly higher levels of 15 N 2 accumulated in the roots and shoots of plants inoculated with the mutant strain, which indicated that more fixed N was readily available for root uptake and assimilation. Roots and shoots of uninoculated plants exposed to 15 N 2 had a small, consistent but nonsignificant increase in levels of 15 N compared with treatments that were exposed only to the natural atmospheric abundance of the isotope. These results show that the NH 4 + -excreting mutant strain of A. variabilis has the potential to increase N input for plant growth in rice production systems.

An assay was developed which identifies individual bacterial tumbles and so allows rapid, quantitative measurements of tumble frequency in free-swimming bacteria. Tumble frequency is modulated by cells to enable chemotaxis. Mutations in the chemotaxis signal transduction pathway typically have phenotypes of altered tumble frequency. The purpose of this assay is to quantitatively measure steady-state tumble frequency to enable comparisons of mutant strain phenotypes. It was developed using Escherichia coli but should be applicable to other species with a peritrichous flagellation pattern, such as Salmonella typhimurium. Tumbles are defined by a combination of the parameters rate of change of direction and swimming speed, with a rapid change of direction defining the beginning of a tumble and increased swimming speed defining the end. These parameters have previously been shown to be correlated with tumbles in general but not used to identify discrete tumble events. The computer assay was validated by comparing its results with manual observations by eye. The assay was intended to be most sensitive to swimming patterns similar to wild type so as to resolve subtle changes which would result from partial-function mutations. It quantitatively detects extreme behavioral phenotypes as well and can be modified to increase resolution at either extreme if necessary.

Ten lactating goats were randomly assigned to two groups of five vaccinates and five controls. The vaccine was a heat-killed rough mutant strain of Escherichia coli (E. coli) J5 containing 1.5 × 109 organisms per ml in incomplete Freund's adjuvant. The vaccinated group received three 1 ml doses of vaccine 14 days apart. The control group received three 1 ml doses of incomplete Freund's adjuvant in a similar regimen. Seventeen days after the third dose of vaccine or Freund's adjuvant all goats were intramammarily (IM) challenged with 15 colony forming units of live virulent E. coli into each teat. Severe clinical signs such as depression, anorexia and prolonged recumbency were observed after challenge in the control goats. In vaccinated goats the signs were less severe, although they had some depression and anorexia for 36 h postchallenge. Body temperature of the animals in both groups increased after challenge with live E. coli. However, peak temperature was significantly lower in vaccinated goats compared with control goats. E. coli count in the milk of vaccinated goats was significantly lower compared with control goats after IM challenge. E. coli from the milk of vaccinated goats was removed more rapidly compared with control goats. Serum J5 antibody levels of vaccinated goats were significantly higher on Days 14, 21, 28, 35 and 42 after vaccination. After challenge a significantly higher serum J5 antibody level was observed in vaccinated goats at 12, 24, 48 and 72 h. Milk anti-J5 antibody levels were significantly higher on Day 28 after vaccination in vaccinated goats. This study concludes that J5 vaccine provides partial protection against E. coli intramammary challenge and goats can be used as an economic model for studying bovine mastitis.

Mutations in Salmonella typhimurium fliG, fliM and fliN give rise either to non-flagellate, non-motile or non-chemotactic mutant bacteria. The FliG, FliM and FliN proteins form part of recently characterized extended flagellar basal structures, and have been postulated to form a mutually interacting structural complex. We have examined basal body preparations from non-motile or non-chemotactic fliG, fliM and fliN mutant strains by electron microscopy and immunoblot gel analysis. Most flagellar preparations isolated from the non-motile mutants lacked FliM, but contained FliG. The basal bodies lacked the belled morphology characteristic of the wild-type structures, but had protrusions which could be labelled with anti-FliG. Non-motile mutant preparations severely depleted of FliG but containing FliM were also obtained. These preparations contained extended, belled flagellar structures that were labelled with anti-FliM. Thus, FliM is part of the shell of the extended structures responsible for the belled morphology, while FliG may be part of the inner substructure. The extended basal structures from a FliG temperature-sensitive mutant strain rapidly lost FliM, as well as FliG, upon a shift to a non-permissive temperature, implying interaction between the FliG- and FliM-containing substructures. In dramatic contrast to non-motile mutants, extended basal structures isolated from non-chemotactic mutants were indistinguishable from wild-type structures. This difference may reflect the energetics of the different protein-protein interactions operative during torque generation and the switching of rotation sense.

A real-time automated method was developed for simultaneous measurements of phototactic orientation (phototaxis) and step-up photophobic response of flagellated microorganisms. Addition of all-trans retinal restored both photoresponses in a carotenoid-deficient mutant strain of Chlamydomonas reinhardtii in a dose-dependent manner. The phototactic orientation was biphasic with respect to both the light intensity and the concentration of retinal. All-trans retinal was more effective than 11-cis retinal to regenerate both photobehavioral responses. Analogs having locked 11-cis configurations and a phenyl ring in the side chain also induced photoresponses, although at concentrations more than two orders of magnitude higher than all-trans retinal. According to the present assay method, the responses were hardly detectable in cells incubated with retinal analogs in which the 13-ene was locked in either its trans or cis configuration. The results strongly suggest that the isomerization of the 13–14 double bond is important for photobehavioral signal transduction and that a single retinal-dependent photoreceptor controls both phototactic and photophobic responses.

Homoserine dehydrogenase (E.C. 1.1.1.3) from a mutant strain of Corynebacterium glutamicum was purified using the pseudo-specific affinity dye Cibacron Blue F3G-A. The triazine dye was coupled either directly or via 1,4-diaminobutane to Sepharose 4B. Both the recovery and purification factor were significantly higher when the dye was coupled to the gel matrix via the spacer. Using this configuration, a recovery of about 62% was obtained with a purification factor of about 95-fold, achieved in a rapid one-step protocol.

A sensitive procedure for the determination of nitrate within the 1- to 50-microM range is described. The method is based on the photoreduction of nitrate to nitrite by whole cells of a nitrite reductase-less mutant strain of the unicellular cyanobacterium Synechococcus R2. Cell suspensions of this cyanobacterium retain high levels of nitrate photoreduction activity for at least 2 months when maintained in the presence of dimethyl sulfoxide at -70 degrees C.

Necrotizing fasciitis, according to a recent warning circulating on the Internet, can be caught from Costa Rican bananas. But don't believe the hype, says the CDC. The story is unfounded and, like many others, is deconstructed on a CDC website devoted to hoaxes and rumors. The site provides accurate information in the face of some terrifying tales about various contagious diseases. For example, the public was recently alarmed by a claim, apparently spread by the media, that the CDC had discovered a mutant strain of HIV that could be transmitted through the air. The CDC has corrected this misinformation. Luckily, neither HIV nor necrotizing fasciitis is spread as easily as the rumours about them. AVhttp://www.cdc.gov/hoax_rumors.htm

Structures 1 , 2 and 3 have been assigned to rifamycins P, Q and Verde, novel metabolites isolated from a mutant strain of Nocardia mediterranea both on the basis of spectroscopic evidence (UV, IR, MS, 1 H and 13 C NMR) in comparison with the model compounds rifamycin S ( 4 ), rifampicin ( 5 ) and 4-dimethylamino-4-deoxy-rifamycin SV ( 6 ), and of an unambiguous synthesis from rifamycin S ( 4 ).

Using E. coli tRNA precursors isolated from an RNAase P mutant strain, we have studied the steps required for the formation of tRNAs having a mature primary sequence in vitro. Our results suggest that at least three different enzymatic activities can participate in the processing of tRNA precursors.

1. 1. Xanthurenic acid, 3-hydroxykynurenine, and kynurenine hydroxylase activity in bw, st and bwst of D. melanogaster were determined. 2. 2. Both pupa and adult of bw, st and bwst showed much the same level of kynurenine hydroxylase activity. 3. 3. The pupae of bw, st and bwst contained not a little 3-hydroxykynurenine. In st and bwst , the 3-hydroxykynurenine rapidly decreased at emergence, while in bw , the decrease was rather gradual. 4. 4. In bw and st , a considerable amount of xanthurenic acid was synthesized during the pupal stage, while, in bwst , the synthesis was moderate. Moreover, in the adult stage, xanthurenic acid was much less in bwst than in bw and st .

A convenient method for the site-specific cloning of chromosomal genes of Escherichia coli to multicopy plasmids is described. In this scheme a ColEl-derived plasmid containing a terminal fragment of the gene to be cloned (e.g, from a gene fusion) is first integrated into the chromosome of a PolA− mutant strain. The plasmid integrate is then rescued in such a way that the intact copy of the gene is included. We demonstrate the applicability of this method by cloning the wild-type ompC gene of E. coli.

A mutant of Saccharomyces cerevisiae has been obtained that shows an absolute growth requirement for long chain bases found in sphingolipids. In the absence of a long chain base, the cells are unable to synthesize the phosphoinositol-containing sphingolipids characteristic of yeast. These results suggest that one or more of the yeast sphingolipids plays a vital biological role.

Experience-dependent modification of courtship behaviour in Drosophila melanogaster has been previously reported; males that have courted fertilized females thereafter avoid courtship with virgin females for 2–3 h. We report that this modification is dependent upon a behavioural interaction between a courting male and fertilized female. Behavioural comparisons of males of the mutant strain Don Giovanni (dg) and normal males imply that fertilized females release a conditioning cue in response to male courtship. (1) The normal behaviour modifying capacity of fertilized females is dependent upon copulation. Females fertilized by dg do not effectively modify male courtship. (2) dg males are deficient in causing fertilized females to release a conditioning cue; fertilized females which are courted by dg males do not induce modification of behaviour. (3) The behaviour of dg males can be modified if the fertilized females they court have been first courted by normal males. These defects were mapped to the y-cv interval of the X-chromosome.

The structure of the oligosaccharide portion of an E.coli Re lipopolysaccharide was determined as α-KDO-(2→4)-α-KDO-(2→6)-β-GlcN-α(1→6)-α-GlcN, bisphosphorylated at positions 1 and 4′. Taking into account the previous determination of the acylation pattern of the GlcN disaccharide, the total structure of E.coli Re LPS was thus established.

Four enzymic components, coded for by the entD, entE, entF and entG genes, involved in the biosynthesis of enterochelin from 2,3-dihydroxybenzoate have been separated from cell extracts of mutant strains of Escherichia coli K-12. The starting material for fractionation of the E, F and G components was a cell extract of an entD mutant strain, which yielded the E, F and G enzymic components uncontaminated by a functional D component. The D component was isolated from cell extracts of an entE mutant strain. The conversion of 2,3-dihydroxybenzoate and l-serine into enterochelin is dependent on the presence of all four enzymic components. The E and F components were shown to catalyze ATP-pyrophosphate exchange reactions dependent on 2,3-dihydroxybenzoate and l-serine, respectively, whereas fractionated extracts of the entE and entF mutant strains lacked these reactions. These data provide firm evidence that the E and F components are involved in the initial activation of the substrates. The D and G components are necessary for subsequent and, as yet, undefined reactions.

The capsular polysaccharides from Cryptococcus neoformans serotype A and from a possible, mutant strain thereof have been structurally analyzed. The latter material is similar to the capsular antigen of C. neoformans serotype D.

Cultures ofAspergillus parasiticus produce the polyketide versicolorin A in response to elevation of the Zn2+ content of the growth medium. With suboptimal Zn2+ (0.8 μM) mycelial growth is about half maximal, and versicolorin synthesis is essentially zero. Inclusion of Cd2+ (1–100 μM) in the Zn2+-limiting growth medium allows optimal growth and stimulates full versicolorin synthesis. Cd2+, like Zn2+, will stimulate versicolorin sysnthesis only when added within the first 30 h after conidial inoculation. The transport system for Cd2+ uptake may be the same as that for Zn2+, as judged byin vivo competition studies. Cd2+ is a competitive inhibitor of Zn2+ uptake, with Ki = 20 μM.

A fraction of UVA-induced 8-methoxypsoralen (8-MOP) mono-adducts can be transformed be a second UVA (365 nm) irradiation procedure into lethal cross-links in Sacccharomyces cerevisiae . To follow the fate of cross-linkable mono-adducts, cells were incubated in complete medium between the two UVA doses and survival was measured. The killing effect of the second UVA dose decrease rapidly in haploid wild-type as well as in strains blocked in mutagenic ( RAD6 + type) or in recombinogenic ( RAD52 + type) repair pathways. This is also true in the pso1-1 and pso2-1 strains selected for sensitivity to 8-MOP plus UVA treatment. In contrast, persistence of mono-adducts is observed in strains blocked in the excision-resynthesis repair pathway. In other words, cross-linkable mono-adducts are repaired by the excision process. The use of the cell-cycle conditional mutant strain ( cdc14-1 ) permitted us to applu the second dose at a specific cell-cycle stage (post-G2 phase) after a ‘priming’ UVA treatment on stationary (G1) phase cells. Such experiments showed a bypass of mono-adducts in an excision-deficient context for at least one round of DNA replication.

We have examined the time and location of the first septa formed by cells of Candida albicans when such cells were induced to grow synchronously as either mycelial or budding cultures. Synchronous growth was initiated by inoculating stationary-phase cells into an amino acid medium with a pH of either 6.5 or 4.5. Septa were visualized with the fluorescent stain Calcofluor. Evidence is presented that: (1) During both synchronous mycelium and bud formation, lightly staining septa first appear and 30 to 40 minutes later abruptly convert to darkly staining septa. (2) Although the times of evagination for mycelium and bud formation are the same, the times of septum formation differ. During bud formation the average septum forms very close to the time of initial evagination; during mycelium formation the average septum forms 30 minutes after evagination. (3) The location of septum formation differs. During bud formation, the septa invariably form at the junction of the mother cell and bud; during mycelium formation, the average septum forms in the tube approximately 2 μm from the junction of mother cell and tube. (4) During mycelium formation, the average septum forms when the mycelium is approximately 7 μm in length, and no tube elongation occurs between septum and mother cell once the initial septum is formed. A mutant strain, M11, is also described which begins to form a mycelium under conditions conducive to mycelium formation, but upon septation generates a bud distal to the septum.

Supplementation of cultures ofAspergillus parasiticus with Zn2+ stimulates the synthesis of versicolorin A only if the supplemental Zn2+ is present between 20 and 30 h postinoculation, during early vegetative growth. Cultures which are grown with minimal Zn2+ avidly internalize Zn2+ which is added late in the growth phase or in early stationary phase. A 15-min exposure to 10 μM Zn2+ during the responsive period, 20–30 h postinoculation, prevents this later uptake of Zn2+. The Zn2+ content of the mycelia at the end of the responsive period, 30 h postinoculation, in some way determines subsequent metabolism of the organism. Versicolorin synthesis, which begins about 50 h postinoculation, is directly proportional to the Zn2+ content of the mycelia at 30 h. The uptake of Zn2+, measured late in the growth phase or in early stationary phase, is inversely proportional to the Zn2+ content of the mycelia at 30 h.

The role of complement in the bactericidal activity of human serum against a mutant strain of Pseudomonas aeruginosa used as a model was demonstrated. The involvement of complement in the bacterial destruction of P. aeruginosa, and the contribution of the alternative and classical pathways of the complement system were directly evidenced by using sera from complement-deficient patients.

A mutant strain (PL pT 1143) of Pseudomonasputida PL, has been isolated for its inability to growth with p-cymene as carbon source. The mutant oxidizes p-cymene (and p-cumate) to a compound (λmax 293 nm) which is readily converted to 3-hydroxy-p-cumate by acid. 4-Trifluoromethylbenzoate is oxidized by the mutant to an acid-stable intermediate (λmax 277nm) that has been crystallized. The spectral properties (u.v., i.r., NMR and mass) of this metabolite are consistent with those expected for a 2,3-dihydro-2,3-dihydroxy derivative of 4-trifluoromethylbenzoate. Further support of this structure was provided by elemental analysis and the properties of two derivatives of the metabolite, 4-trifluoromethyl-3-hydroxybenzoate and an acetonide formed with 2,2-dimethoxypropane. The stability of a product obtained by treatment of the dihydrodiol metabolite with triacetylosmate indicates that it is the cis-isomer.

On the basis of spectroscopic studies (UV, IR, MS, 1H and 13C NMR) on rifamycin W and its tetrahydro- and dimethyl-derivatives, in comparison with the model compound rifamycin S (2), structure 1 is assigned to rifamycin W, a novel ansamycin isolated from a mutant strain of Nocardia mediterranei. The structural similarity of rifamycin W with other rifamycins and with streptovaricins is discussed from the biosynthetic point of view.

Incorporations of singly and doubly-labelled acetate-[13C] into griseofulvin by a mutant strain of Penicillium patulum confirm its origin from simple folding of a single heptaketide chain. An acetate ‘starter’ effect is observed in the 13C-NMR spectra of griseofulvin enriched from acetate-[13C], and analysis of the 13C—13C spin—spin couplings observed indicate a rapid metabolic turnover of added acetate. Methyl, but not carboxyl, of acetate is efficiently metabolised into the C1 pool.

A rapid method for quantification of versicolorin, based upon the red shift in the visible absorbance spectrum upon ionization of the phenolic hydroxyl groups, has been developed. The production of versicolorin requires zinc concentrations in excess of those required for growth. Maximal growth occurs with 2 μM zinc; maximal versicolorin production requires 5 μM zinc in the medium. Maximum production of versicolorin occurs after cessation of rapid growth, but does not absolutely depend upon exhaustion of the carbon source.

2,3-Dihydroxy- N -benzoylserine was synthesized. A number of its properties were compared and found to be identical with those of a natural product of formed by Escherichia coli and Aerobacter aerogenes . The synthetic 2,3-hydroxy- N -benzoylserine acts as a growth factor for a mutant strain of E. coli .