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In the present investigation, significant physical parameters such as incubation time, temperature and pH were optimized for cellulase production by wild and mutant strains of thermophilic Thermomyces dupontii. For a comparative analysis, three independent variables which were selected from OVAT were subjected to RSM. As per response surface methodology (RSM) model, central composite design was used in order to analyze the interactive effects of significant physical factors on cellulase productivity. The second polynomial regression model with a determination coefficient R2 value 0.9503 and 0.9711 was found to be adequate and significant. 3D response surface plots and contours showed interaction among three variables. In contrast to wild T. dupontii, mutant strain showed 1.7-fold increase in FPase activity (43 U/ml/min) at 55 °C and pH 5.5 in 72 h. The result indicates thermophilic mutant T. dupontii has a potential for industrial use, because enzyme obtained from thermophile is more appropriate for industrial process where high temperature is involved.

Astaxanthin is widely used in food, feed and nutraceutical industries. Xanthophyllomyces dendrorhous is one of the most promising natural sources of astaxanthin. However, the astaxanthin yield in the wild-type X. dendrorhous is considered low for industrial application. In the present study, X. dendrorhous ATCC 66272 was subjected to two-staged mutagenesis: (i) UV light and (ii) N-methyl-N′-nitro-N-nitroso-guanidine (NTG) toward attaining higher astaxanthin yield. The UV-irradiation mutant, X. dendrorhous SK974 showed 1.7-fold (1.07 mg/g) higher astaxanthin production as compared with the wild-type strain (0.65 mg/g). The UV mutant strain was then treated with NTG, designated as X. dendrorhous SK984, displayed further 1.4-fold (1.45 mg/g) higher astaxanthin production. Furthermore, the oak leaf extract (5%, v/v) and inorganic phosphate (KH2PO4, 3 mM) supplementation resulted about 1.4-fold (1.98 mg/g) higher astaxanthin production as compared with control (1.45 mg/g) in X. dendrorhous SK984. These findings serve as a platform suggesting that intersecting approaches might be aimed toward systematically enhanced astaxanthin production.

The SARS-CoV-2 lineage carrying the amino acid change D614G has become the dominant variant in the global COVID-19 pandemic. The rapid spread of the G614 mutant suggests that it may have a transmission advantage over the D614 wildtype. Using our previous epidemiological framework to analyze COVID-19 surveillance and sequence data, we estimated that the G614 mutant is 31% (28-34%) more transmissible than the D614 wildtype. As such, interventions that were previously effective in containing or mitigating the D614 wildtype (e.g. in China, Vietnam, Thailand, etc.) might be less effective against the G614 mutant. Our framework can be readily integrated into current COVID-19 surveillance to monitor the emergence and fitness of mutant strains, such that pandemic surveillance, disease control and development of treatment and vaccines can be adjusted dynamically.

A multiblocked mutant strain (ΔAHBA and Δasm12, asm21) of Actinosynnema pretiosum, the producer of the highly toxic maytansinoid ansamitocin, has been used for the mutasynthetic production of new proansamitocin derivatives. The use of mutant strains that are blocked in the biosynthesis of an early building block as well as in the expression of two tailoring enzymes broadens the scope of chemo‐biosynthetic access to new maytansinoids. Remarkably, a ring‐expanded macrolactone derived from ansamitocin was created for the first time., Make it larger! A new variant of the chemo‐biosynthetic concept of mutasynthesis using a multiblocked strain of the ansamitocin producer A. pretiosum yielded new macrolactam proansamitocins as well as the first ring‐expanded maytansin macrolactone.

Docosahexaenoic acid (DHA), a n-3 long-chain polyunsaturated fatty acid, is critical for physiological activities of the human body. Marine eukaryote Aurantiochytrium sp. is considered a promising source for DHA production. Mutational studies have shown that ultraviolet (UV) irradiation (50 W, 30 s) could be utilized as a breeding strategy for obtaining high-yield DHA-producing Aurantiochytrium sp. After UV irradiation (50 W, 30 s), the mutant strain X2 which shows enhanced lipid (1.79-fold, 1417.37 mg/L) and DHA (1.90-fold, 624.93 mg/L) production, was selected from the wild Aurantiochytrium sp. Instead of eicosapentaenoic acid (EPA), 9.07% of docosapentaenoic acid (DPA) was observed in the mutant strain X2. The comparative transcriptomic analysis showed that in both wild type and mutant strain, the fatty acid synthesis (FAS) pathway was incomplete with key desaturases, but genes related to the polyketide synthase (PKS) pathway were observed. Results presented that mRNA expression levels of CoAT, AT, ER, DH, and MT down-regulated in wild type but up-regulated in mutant strain X2, corresponding to the increased intercellular DHA accumulation. These findings indicated that CoAT, AT, ER, DH, and MT can be exploited for high DHA yields in Aurantiochytrium.

Spontaneous mutations are important players in evolution. Nevertheless, there is a paucity of information about the mutagenic processes operating in most bacterial species. In this work, we implemented two forward mutational markers for studies in Caulobacter crescentus. We confirmed previous results in which A:T → G:C transitions are the most prevalent type of spontaneous base substitutions in this organism, although there is considerable deviation from this trend in one of the loci analyzed. We also investigated the role of dinB and imuC, encoding error-prone DNA polymerases, in spontaneous mutagenesis in this GC-rich organism. Both dinB and imuC mutant strains show comparable mutation rates to the parental strain. Nevertheless, both strains show differences in the base substitution patterns, and the dinB mutant strain shows a striking reduction in the number of spontaneous -1 deletions and an increase in C:G → T:A transitions in both assays.

Aims Yeasts produce 2-phenylethanol (2-PE) from sugars via de novo synthesis; however, its synthesis is limited due to feedback inhibition on the isofunctional 3-deoxy-d-arabino-heptulosonate-7-phosphate (DAHP) synthases (Aro3p and Aro4p). This work aimed to select Kluyveromyces marxianus mutant strains with improved capacity to produce 2-PE from sugars. Methods and results Kluyveromyces marxianus CCT 7735 mutant strains were selected from UV irradiation coupled with screening of p-fluoro-dl-phenylalanine (PFP) tolerant strains on culture medium without l-Phe addition. Most of them produced 2-PE titres higher than the parental strain and the Km_PFP41 mutant strain stood out for displaying the highest 2-PE specific production rate. Moreover it showed higher activity of DAHP synthase than the parental strain. We sequenced both ARO3 and ARO4 genes of Km_PFP41 mutant and identified mutations in ARO4 which caused changes in both size and conformation of the Aro4p. These changes seem to be associated with the enhanced activity of DAHP synthase and improved production of 2-PE exhibited by that mutant strain. Conclusions The Km_PFP41 mutant strain presented improved 2-PE production via de novo synthesis and enhanced DAHP synthase activity. Significance and impact of the study The mutant strain obtained in this work may be exploited as a yeast cell factory for high-level synthesis of 2-PE.

Currently, algal fuel research has commenced to shift toward genetically engineered mutants able to express and excrete desired products directly into the culture. In this study, a mutant strain of Chlamydomonas reinhardtii, engineered for bisabolene (alternative biodiesel) excretion, was cultivated at different illumination and temperatures to investigate their effects on cell growth and bisabolene production. Moreover, a kinetic model was constructed to identify the desirable conditions for biofuel synthesis. Three original contributions were concluded. First, this work confirmed that bisabolene was partially synthesized independently of biomass growth, indicating its feasibility for continuous production. Second, it was found that while bisabolene synthesis was independent of light intensity, it was strongly affected by temperature, resulting in conflicting desirable conditions for cell growth and product synthesis. Finally, through model prediction, optimal operating conditions were identified for mutant growth and bisabolene synthesis. This study therefore paves the way toward chemostat production and process scale-up.

Previously, activation of the cryptic nam gene cluster led to three new naphthalenic ansamycins with unprecedented n-pentyl and n-butyl side chains from the mutant Streptomyces sp. SR201nam1OE strain. In this study, we further characterized the products of the mutant strain and six new neoansamycin congeners, namely neoansamycins D–I (1–6), were elucidated. Among them, compounds 1–3 feature the conserved skeleton of neoansamycins B and C but with n-hexyl side chains, and 4–6 are modified neoansamycins with n-pentyl side chains, illustrating the biosynthetic plasticity and diverse post-PKS modifications of neoansamycins.

A combination of microbial strain improvement and statistical optimization is investigated to maximize pneumocandin B0 production from Glarea lozoyensis ATCC 74030. Atmospheric and room temperature plasma (ARTP) was used to enhance G. lozoyensis ATCC 74030 in pneumocandin B0 yield. Mutant strain G. lozoyensis Q1 exhibited 1.39-fold increase in pneumocandin B0 production to 1134 mg/L when compared with the parent strain (810 mg/L). Further, the optimized medium provided another 1.65-fold in final pneumocandin B0 concentration to 1873 mg/L compared to the original medium. The results of this study indicated the combined application of a classical mutation and medium optimization can improve effectively pneumocandin B0 production from G. lozoyensis and could be a tool to improve other secondary metabolites production by fungal strains.

Three rare carbamate-containing alkaloids, penicimutamides A–C (1–3), were isolated from a fungal mutant from the diethyl sulfate (DES) mutagenesis of marine-derived Penicillium purpurogenum G59. Their structures, including their absolute configurations, were determined by spectroscopic methods, especially the X-ray crystallography and CD analyses. HPLC-UV and HPLC-MS analyses evidenced that 1–3 were only produced in the mutant strain via biosynthetic pathways that were silent in the parental strain and activated by DES mutagenesis.

Protein-protein interactions are the basis for all biochemical cellular activities. The Ras Recruitment System, RRS, is a method for studying interactions between known proteins as well as identification of novel interactions following a cDNA library screen. The method is based on the recruitment of the Ras protein to the plasma membrane via protein-protein interactions. The interaction between proteins is studied in a temperature-sensitive yeast Saccharomyces cerevisiae mutant strain. This mutant is able to grow under restrictive temperature conditions when the Ras viability pathway becomes activated as a result of a positive protein-protein interaction. The RRS complements the limitations and problems that arise from the yeast two-hybrid system.

A mutant strain of Clostridium beijerinckii, with high tolerance to ferulic acid, was generated using atmospheric pressure glow discharge and high-throughput screening of C. beijerinckii NCIMB 8052. The mutant strain M11 produced 7.24 g/L of butanol when grown in P2 medium containing 30 g/L of glucose and 0.5 g/L of ferulic acid, which is comparable to the production from non-ferulic acid cultures (8.11 g/L of butanol). When 0.8 g/L of ferulic acid was introduced into the P2 medium, C. beijerinckii M11 grew well and produced 4.91 g/L of butanol. Both cell growth and butanol production of C. beijerinckii M11 were seriously inhibited when 0.9 g/L of ferulic acid was added into the P2 medium. Furthermore, C. beijerinckii M11 could produce 6.13 g/L of butanol using non-detoxified hemicellulosic hydrolysate from diluted sulfuric acid-treated corn fiber (SAHHC) as the carbon source. These results demonstrate that C. beijerinckii M11 has a high ferulic acid tolerance and is able to use non-detoxified SAHHC for butanol production.

Caenorhabditis elegans postembryonic development consists of four discrete larval stages separated by molts. Typically, the speed of progression through these larval stages is investigated by visual inspection of the molting process. Here, we describe an automated method to monitor the timing of these discrete phases of C. elegans maturation, from the first larval stage through adulthood, using bioluminescence. The method was validated with a lin-42 mutant strain that shows delayed development relative to wild-type animals and with a daf-2 mutant that shows an extended second larval stage. This new method is inherently high throughput and will finally allow dissecting the molecular machinery governing the speed of the developmental clock, which has so far been hampered by the lack of a method suitable for genetic screens., Our work is supported by the Ludwig-Maximilians-University Munich and the Friederich-Bauer-Stiftung (M.M., M.O., and M.G.); the European Research Council (ERC-2011-StG-281691) and the Spanish Ministerio de Economía y Competitividad (BFU2012-35509) (M.A.-S.); and a Marie-Curie Intra-European Fellowship (FP7-PEOPLE-2013-IEF/GA Nr: 627263) (to M.O. and M.A.-S.).

Supplementing Bacillus (B.) subtilis mutants selected to overproduce a specific amino acid (AA) may be an alternative method to provide essential AA in pig diets. Two experiments on a B. subtilis strain selected to overproduce Trp were conducted using 8-kg pigs fed Trp-deficient diets for 20 d. B. subtilis were supplied in a low or high dose in Experiments 1 and 2, respectively. The Trp-deficient diet (0.15 SID Trp:Lys) reduced (p

In this study, a microalgae mutant Scenedesmus sp. Z-4 with a lipid content of 28.86% and biomass of 2.876 g L−1 was obtained using 60Co γ-ray mutation. The lipid productivity (138 mg L−1 per day) and content of mutant Z-4 were enhanced by 113% and 71.3% compared to that of the wild strain, respectively. In addition, unlike the wild strain, the microalgae cells were larger and the surface was rougher in mutant Z-4.

The use of polygalacturonase (PG) preparations in winemaking promotes the release of phenolic compounds. A PG from a new source, Aspergillus sojae mutant, was semi-purified and tested for grape mash maceration. Crude extract (CE), a commercial pectinase, and two high PG activity semi-purified preparations, FI and FII , were applied for maceration at PG activity of 3.5 U g−1 of grape for 46 h. Enzyme-assisted maceration significantly (P < 0.05) increased the total phenolic content from 255.8 to 916.3 ± 5.2, 5732.9 ± 9.9, 563.4 ± 6.7 and 620.6 ± 18.4 mg L−1 for CE, commercial pectinase, FI and FII, respectively. The content of individual phenolics such as gallic, protocatechuic, chlorogenic and p-coumaric acids was improved. Principal component and hierarchical clustering analyses suggested that CE has a better performance upon the release of phenols. Semi-purified preparations acted similar to commercial pectinase. These findings open an opportunity for the potential use of PG from the mutant strain as an alternative macerating enzyme., EU-FP7-Marie Curie Action (PIRSES-GA 2010-269211); Biotechnology and Synthetic Biology Focus Group (0020DII001 0820000100); National Council on Science and Technology of Mexico (CONACyT) (204152)

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.

The R. glutinis NR98 of high-producing β-carotene was treated by ultra high pressure (UHP) of 100~500 MPa for 10~30 min. The survival curve of NR98 treated for 10 min was saddle, which shows that the effect of UHP on this strain maybe was the cumulation of many biology effects. The rate of β-carotene production from mutant G39 (obtained at 300 MPa for 10 min) was increased by 59.87% compared with the initial strain NR98, and its genetic quality was proved to be stable by experiments. The result of restriction fragment length polymorphism analysis suggested that mutant strain G39 was likely to change in nucleic acid level.

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 high molecular weight hyaluronic acid-producing mutant, designated asStreptococcus equiSH-109 that did not produce streptolysin or hyaluronidase was selected from wild-typeStreptococcus equiSH-0 by NTG treatment. The selected mutant could produce high molecular weight HA and had perfect genetic stability after reproduction. Several culture conditions were optimized in a 5-l fermentor for the production of hyaluronic acid. It was found that the optimal pH and agitation speed were 7.6 and 550rpm, respectively. Under the optimized culture condition, hyaluronic acid with a molecular weight of 4.0 x 106was accumulated in the culture broth.

Supplementing a culture of a mutant strain of Actinosynnema pretiosum that is unable to biosynthesize aminohydroxy benzoic acid (AHBA), with 3-azido-5-hydroxy-benzoic acid and 3-azido-5-amino-benzoic acid, unexpectedly yielded anilino ansamitocins instead of the expected azido derivatives. This is the first example of the bioreduction of organic azides. The unique nature of these results was demonstrated when 3-azido-5-amino-benzoic acid was fed to the corresponding AHBA blocked mutant of Streptomyces hygroscopicus, the geldanamycin producer. This mutasynthetic experiment yielded the fully processed azido derivative of geldanamycin.

wild type strain W3110; in the YS1234 strain defective in ynbD, yeiU, ybjG, and pgpB; in the YS1235 strain defective in ynbD, yeiU, ybjG, and bacA; and in the YS1245 strain defective in ynbD, yeiU, pgpB, and bacA were 78.0% ± 3.5%, 74.7% ± 1.2%, 68.4% ± 6.8%, and 64.0% ± 7.5%, respectively. The quadruple mutant strain that possessed the wild-type bacA showed almost identical UP level as the wild type strain W3110, indicating that sufficient level of UPP phosphatse activity was conferred by a bacA product without the products of ynbD, yeiU, ybjG, and pgpB. However, the mutants defective in bacA showed a slight but significant (p < 0.01) decrease in the UP level.

Three rare carbamate-containing alkaloids, penicimutamides A–C (1–3), were isolated from a fungal mutant from the diethyl sulfate (DES) mutagenesis of marine-derived Penicillium purpurogenum G59. Their structures, including their absolute configurations, were determined by spectroscopic methods, especially the X-ray crystallography and CD analyses. HPLC-UV and HPLC-MS analyses evidenced that 1–3 were only produced in the mutant strain via biosynthetic pathways that were silent in the parental strain and activated by DES mutagenesis.

IS-Tlk/Kyo, a rat mutant strain derived from IS/Kyo strain, exhibits a kinked and/or short tail, in addition to a congenital anomaly of the lumbar vertebrae that is a hallmark of IS/Kyo rats. Homozygotes (Tlk/Tlk) of Tlk dominant gene are known to die during embryonic development. The present report deals with the morphological features of heterozygous IS-Tlk/Kyo rat fetuses in comparison with those of IS/Kyo rat fetuses. One of the morphological features was a high incidence of tail vertebral anomalies in IS-Tlk rats (81.6% versus 0% in IS/Kyo rats). Significantly low values in number of live fetuses and ossified 5th sternebra and sacral and caudal vertebrae were observed in IS-Tlk/Kyo rats compared with those in IS/Kyo rats as well as a low incidence of fetuses with ventral septal defects in IS-Tlk/Kyo (0% versus 54.4% in IS rats). These results suggest that the Tlk gene may be involved in the formation of the vertebral centra and the ventral septum when it expresses on the genetic background of the IS rat.

The aim of the study was to evaluate baseline data of mutant strain of mice and to compare haematological parameters with their background strains. Almost all mutant strains of either sex had statistically significant changes in relation to their background strains. These findings will be useful for researchers in designing experiments on mutant strains for various disease models and interpreting data obtained from those strains.

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.

A fundamental possibility of antitumor activity testing in the bacterial system represented by Escherichia coli of the wild type and its MS2-induced mutant has been shown. The initial bacterial strain is an indicator of toxic properties of tested substances and the mutant strain is a specific test culture modeling a tumor cell. The comparison of the data described for eukaryotes with the data obtained using the proposed bacterial test system confirms an adequate response of both strains to substances with proved antitumor properties. The data obtained are considered as very promising for the further improvement of this test system towards its application for primary screening of antitumor substances.

It has been suggested that Vibrio vulnificus attaches to plankton and algae and is found in large numbers in the environment. Factors affecting attachment, biofilm formation and morphology of V. vulnificus have not been thoroughly investigated. This study evaluated the role of quorum sensing (QS) and environmental conditions on biofilm development of V. vulnificus. It was found that biofilm development by V. vulnificus was affected by nutrient and glucose concentration, but not by NaCl concentration or temperature under the conditions used here. Moreover, biofilm development of a QS mutant strain proceeded rapidly and sloughing occurred earlier than for the isogenic parent strain. There was a significant loss of viability for the QS mutant biofilm early in development. Hence, it is hypothesised that factors regulated by the QS system play a role in proper biofilm development and maintenance of V. vulnificus. Furthermore, it is shown that biofilm development varied among isolates.

Three novel polyketide metabolites were isolated from laboratory-scale fermentation of the Streptomyces rimosus mutant strain R1059. Structural elucidation of the compounds was based on NMR experiments. The compounds were characterized as naphthalene derivatives: (rel)-4beta,8-dihydroxy-3alpha-hydroxymethyl-4alpha-methyl-1,2,3,4-tetrahydronaphthalene1-one (1), 4xi8-dihydroxy-3-hydroxymethyl-4xi-methyl-1,4-dihydronaphthalene-1-one (2) and (rel)-4beta,8-dihydroxy-3alpha-O-[alpha-glucopyranosyl]hydroxymethyl-4alpha-methyl-1,2,3,4-tetrahydronaphthalene-1-one (3). The compounds isolated appear to be derived via a shorter polyketide backbone than oxytetracycline (4), the normal end-product made by the parent of this strain. Compound 3 was the glucoside of 1 and must be formed as a post-PKS reaction by the activation of a glycosyl transferase, which has not been reported in this species before.

Strains of Rhizobium spp. isolated from leguminous plants and standard strains accumulated 27% to 57% polyhydroxyalkanoate (PHA) of their cell biomass. Among these cultures, one strain of Rhizobium meliloti synthesized 10-30% more PHA than others and contained 3% hydroxyvalerate (HV) when grown on sucrose as carbon substrate. The occurrence of hydroxybutyrate (HB) and HV was confirmed by GC and 1H NMR analysis. Treatment of the culture with 4'-N-piperidinobutyl-2-chlorophenoxazine resulted in a mutant which synthesized up to 69%, PHA of the cell biomass with an improved yield of 11 to 47% under different carbon and nitrogen ratios, compared to the parent strain.

Two new avemectin analogues 1 and 2 belonging to 'b' series of components were isolated from a ∆aveCDE mutant Streptomyces avermectinius strain. Their structures were elucidated by detailed analysis of NMR and MS spectroscopic data. Compounds 1 and 2 displayed moderate cytotoxicities against B16, MG-63, and Saos-2 cell lines in an in vitro assay.

Several levan hyperproducing mutants of Zymomonas mobilis strains were selected by mutagenesis with N-methyl-N-nitro-nitrosoguanidine and caffeine. Highest levan production (41 g l−1) was obtained with a mutant strain HL 29 in a culture medium containing 200 g sucrose l−1 and 0.5 g (NH4)2SO4 l−1 stored at 7 °C for 29 days. This is the first report describing the levan synthesis by Z. mobilis at 7 °C.

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.

A developmental mutant of A. nidulans strain (MC430) was obtained by NTG treatment. The mutant shows secondary conidiophores, multiple tiers of sterigmata, absence of conidiophore pigmentation and foot-cell alterations. This phenotype was determined by a gene (medA430) mapped on the chromosome I. The map distance between the medA430 and galD loci was 23.7 cM.It's the same location of the medA locus. The foot-cell abnormalities present in this mutant strain indicate that it is a carrier of a new allele of the medA gene. It's very useful for future analysis with regard to the Med protein action on the conidiophore development.

Gnotobiotic piglets inoculated with Escherichia coli O157:H7, its luxS mutant derivative, or nonpathogenic E. coli were evaluated for attaching and effacing lesions. Although no differences in clinical symptoms were seen between pigs inoculated with the parent and those inoculated with the luxS mutant, the luxS mutant-inoculated pigs had a lower frequency of attaching and effacing lesions in the spiral colon than parent strain-inoculated pigs.

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.

The available passive mode of periodontal infections in mice requires high efficiency of bacterial attachment and invasiveness and is not always suitable to test the pathogenicity of genetically engineered mutant strains. We developed an active mode of oral infection, using microinjection in the marginal gingiva of mice, to test the pathogenicity of a genetically engineeredTreponema denticolamutant strain deficient in intermediate-like filaments, compared to the wild-type strain. This targeted mode of infection inoculates the bacterial strain to be tested directly at a lesion site (needle entry point) located at the future periodontal lesion site. The efficiency ofT. denticolawild-type strain to elicit bone loss contrasted with the lack of pathogenicity of the intermediate-like filament deficient mutant strain in comparison to the sham infection. The periodontal microinjection oral model in mice can be used for a variety of applications complementary to the passive mode of periodontal infection in context of pathogenicity testing.

Polyacrylamide gel electrophoresis profiles of various cell surface lipopolysaccharides (LPS) obtained from fast-growing strains and plasmidless mutant strains of Rhizobium ‘hedysari’, R. leguminosarum and from slow-growing Bradyrhizobium japonicum strains have been compared. Correlations were obtained by using serological techniques. Two of the fast-growing strains and one slow-growing strain employed showed several LPS with slow mobility and a few others characterized by fast mobility in the gel. Other fast-growing rhizobium strains and all the other slow-growing bradyrhizobia tested had bands widely distributed in the gels. Location of bands, in these rhizobia, did not correlate with rapidity of growth of the strains. Induced antibiotic resistance did not affect LPS composition for either the fast- or slow-growing rhizobia. The LPS composition was apparently affected by the loss of plasmids. It appears to be associated with the lack of some bands in the region ranging from 30–50 kDa. The LPS composition was also affected by the mutation in the fast mobility region. The mutant strain lacked a low molecular mass band compared with the parental strain. [ 1 H]-NMR spectra of LPS extracts from the latter strain also revealed a signal that is missing from the spectrum of the LPS of mutant strain.

In Salmonella enterica serovar Typhimurium, MgtC and PagC are positively regulated by the PhoP–PhoQ two-component system, which is activated under magnesium deprivation. Both MgtC and PagC are of unknown function but have been involved in intramacrophage survival. We have found that the amount of PagC is lowered in a ΔmgtC mutant strain grown in magnesium depleted medium. However, the effect of MgtC on PagC does not account for the growth defect of a ΔmgtC mutant in macrophages since, in contrast to previous reports, our results indicate that PagC does not contribute to intramacrophage survival. In addition, a pagC null mutant is only poorly attenuated in Nramp1-negative or Nramp1-positive mice. On the other hand, a mgtC null mutant is significantly more attenuated with Nramp1-positive than Nramp1-negative mice, suggesting that a functional Nramp1 (Slc11a1) further limits the multiplication of this mutant within the host.

We have constructed a near-saturation level mutant library for Pseudomonas aeruginosa strain PAO1 using Tn5-derived transposons mapped to the PAO1 reference sequence. This chapter describes the high-throughput techniques used to generate and map the mutant strains. In addition, an analysis of the utility of this collection is presented based on changes to the annotation for the PAO1 genome in the past years, as well as the citation record for this collection. It is clear that many avenues of research have been accelerated by this collection and that additional large mutant strain collections will further aid in defining gene function and biological processes in pathogens.

Pseudomonas testosteroni M4-1, capable of using phthalate as the sole carbon and energy source, was isolated. Tn5 mutagenesis using pSUP2021 yielded mutant strains of M4-1 that are defective in phthalate metabolism and produce a dihydrodiol compound. The dihydrodiol compound produced by mutant strain M4-122 was isolated and identified as 4,5-dihydro-4,5-dihydroxyphthalate (DDP) by elementary analysis, mass analysis and nuclear magnetic resonance. Various conditions to increase the yield of DDP from phthalate were examined for mutant strain M4-122. With resting cells 6 g DDP/1 were produced. The additional of ethanol to the resting-cell reaction mixture enhanced DDP production and 10 g DDP/1 was produced from 8.3 g/1 of phthalate.

After UV-irradiation an alkaloid-blocked mutant of Claviceps paspali was isolated from an ergometrine and lysergic acid amides (1:1) producing strain. The formation of ergometrine is blocked in this particular mutant strain which accumulates only lysergic acid amides. A significant morphological differentation of colonies of mutant as well as of mycelium in submerged cultures could be observed and total alkaloid yields are rather influenced.

Aspergillus niger van Teighem, a soil isolate, was mutagenized by u.v. light. A mutant strain with 3 times increased productivity was generated on exposure of the parent strain to u.v. for 15 min. This mutant on further exposure to u.v. yielded second generation mutants with only marginal increase in inulinase productivity with respect to the parent mutant, but the best yield of 377 U ml-1 was considered promising for industrial use.

The application of high-resolution 1 H, 13 C and 31 P NMR and MS analyses to the oligosaccharide moieties of the L4 and L7 immunotypes of Neisseria meningitidis revealed that they had the following structures: The fact that the L7 LPS is not sialylated at O-3 of its terminal β- d -galactopyranosyl residue implies that it is a mutant strain unable to endogenously sialylate its lacto- N -neotetraose antenna. With the structural elucidation of the L4 and L7 LPS immunotypes, a more comprehensive structural profile of the LPS involved in disease isolates can now be assembled. This provides valuable insights into the structural basis of the N. meningitidis immunotyping system which could be of use in formulating an LPS-based vaccine against meningococcal meningitis.