Alginate beads as a storage, delivery and containment system for genetically modified PCB degrader and PCB biosensor derivatives of Pseudomonas fluorescens F113

Aims: Pseudomonas fluorescens F113Rifpcb is a genetically engineered rhizosphere bacterium with the potential to degrade polychlorinated biphenyls (PCBs). F113Rifpcbgfp and F113L::1180gfp are biosensor strains capable of detecting PCB bioavailability and biodegradation. The aim of this paper is to evaluate the use of alginate beads as a storage, delivery and containment system for use of these strains in PCB contaminated soils. Methods and Results: The survival and release of Ps. fluorescens F113Rifpcb from alginate beads were evaluated. Two Ps. fluorescens F113‐based biosensor strains were encapsulated, and their ability to detect 3‐chlorobenzoate (3‐CBA) and 3‐chlorobiphenyl (3‐CBP) degradation in soil was assessed. After 250 days of storage, 100% recovery of viable F113Rifpcb cells was possible. Amendments to the alginate formulation allowed for the timed release of the inoculant. Encapsulation of the F113Rifpcb cells provided a more targeted approach for the inoculation of plants and resulted in lower inoculum populations in the bulk soil, which may reduce the risk of unintentional spread of these genetically modified micro‐organisms in the environment. Encapsulation of the biosensor strains in alginate beads did not interfere with their ability to detect either 3‐CBA or 3‐CBP degradation. In fact, detection of 3‐CBP degradation was enhanced in encapsulated biosensors. Conclusions: Alginate beads are an effective storage and delivery system for PCB degrading inocula and biosensors. Significance and Impact of the Study: Pseudomonas fluorescens F113Rifpcb and the F113 derivative PCB biosensor strains have excellent potential for detecting and bioremediation of PCB contaminated soils. The alginate bead delivery system could facilitate the application of these strains as biosensors.     

Living immobilized Acetobacter in Ca-alginate in vinegar production: Perliminary study on optimum conditions for immobilization

Summary The optimum conditions forAcetobacter immobilization were investigated. The results show that: 1) the maximum oxygen uptake rate (OURm) and cell release are related to alginate and cell concentration in the gel; 2)different alginate concentration does not affect cell viability, but long storage in CaCl₂ reduces the number of living cells; 3)the double alginate gel layers had no influence on cell viability and on the OURm and prevented cell leakage from the gel matrix.     

海藻酸盐裂解酶研究进展

海藻酸盐裂解酶是一类降解褐藻中海藻酸盐的酶。此酶已经在多种有机体中得到分离。对海藻酸盐裂解酶的生物特性、研究方法及其生物学功能进行了介绍。在酶学特性研究的基础上 ,通过酶解构建新型海藻酸盐多聚物 ,可增强和扩展海藻酸盐裂解酶在工业、农业、医药领域中的应用 ,使其在海藻多糖的高值化应用中发挥重要的作用。概述了海藻酸盐和海藻酸盐裂解酶过去和现在的研究状况 ,展望了海藻酸盐和海藻酸盐裂解酶将来的应用前景。     

Microencapsulation of canine sperm and its preservation at 4 °C

The objective of this study was to develop a preservation method for canine sperm using microencapsulation. Pooled ejaculates from three beagles (Canis familiaris) were extended in egg yolk Tris extender and were encapsulated in gel (alginate only) or polycation (poly-l-lysine membrane bound) microcapsules at 0.75% and 1.0% alginate concentration. In Experiment 1, characteristics of microcapsule and microencapsulated sperm were evaluated during chilling storage for 48 h. Gel microcapsules at 0.75% alginate concentration had a teardrop-like structure with fragility, whereas those at 1.0% alginate had a solid spherical structure. In all groups, diameter of the microcapsules increased with duration of storage (P < 0.05). Alginate concentration did not affect the sperm recovery rate from microcapsules. Total average recovery rate of sperm from polycation microcapsules was lower than that of gel microcapsules (P < 0.05). Progressive motility of polycation microencapsulated sperm and unencapsulated sperm (control) was higher than that of the gel microencapsulated sperm, both at 0.75% and 1.0% alginate concentration (P < 0.05), although viability of sperm was similar among the three groups. In Experiment 2, to evaluate the sperm longevity after chilling storage, sperm were microencapsulated in polycation microcapsules at 1.0% alginate concentration, stored at 4 °C for 0, 1, 4, and 7 d, and then cultured at 38.5 °C for 0, 6, and 24 h. Progressive motility and viability of microencapsulated sperm were higher than those of unencapsulated spermatozoa at 0 to 24 h of culture after 4 and 7 d of chilling storage (P < 0.05). In conclusion, polycation microencapsulation at 1.0% alginate concentration can be successfully applied for chilling storage of canine sperm by maintaining motility and viability for up to 7 d.     

Immobilization in alginate as a technique for the preservation of Bacillus thuringiensis var. israelensis for long-term preservation

Technique for immobilization using sodium alginate as the matrix to preserve Bacillus thuringiensis var. israelensis isolates for long time storage was developed. Two strains of B. thuringiensis var. israelensis viz., VCRC B-17 and WHO standard strain IPS-82 were immobilized in alginate matrix and preserved at 4 degrees C and when tested both were found to have maintained excellent viability and mosquito larvicidal activity for 10 years. Mosquito larvicidal activity of B-17 and IPS-82 alginate beads, in term of LC(50) values before storage was 72.07 ng/ml and 47.07 ng/ml, respectively and after storage at 4 degrees C for a period of 1 to 10 years the values ranged from 69.88 to 73.86 ng/ml with a mean of 72.38 ng/ml and 45.32 to 48.60 ng/ml with a mean of 47.49 ng/ml, respectively. Similarly spore count of the beads of the respective strains was 4.37 x 10(8) and 3.33 x 10(10) CFU/mg before storage. After storage at 4 degrees C for a period of 1 to 10 years the counts of the beads of the respective strains ranged from 4.23 x 10(8) to 4.83 x 10(8) CFU/mg (mean of 4.49 x 10(8) CFU/mg) and 3.2 x 10(10) to 3.87 x 10(10) CFU/mg (mean of 3.54 x 10(10) CFU/mg). The alginate matrix immobilization technique has many advantages over free cells are that they enhance the stability of both spores and toxin against several physicochemical conditions and confer reduced susceptibility to contamination.     

Plant regeneration from alginate-encapsulated shoot tips of Phylianthus amarus schum and thonn, a medicinally important plant species

Summary A method was developed for plant regeneration from alginate-encapsulated shoot tips of Phyllanthus amarus. Shoot tips excised from in vitro proliferated shoots were encapsulated in calcium alginate beads. The best gel complexation was achieved using 3% sodium alginate and 75 mM CaCl₂·2H₂O. Maximum percentage response for conversion of encapsulated shoot tips into plantlets was 90% after 5 wk of culture on Murashige and Skoog (MS) medium without plant growth regulator. The regrowth ability of encapsulated shoot tips was affected by the concentration of sodium alginate, storage duration, and the presence or absence of MS nutrients in calcium alginate beads. Plantlets with well-developed shoot and roots were transferred to pots containing an autoclaved mixture of soilrite and peat moss (1∶1). The conversion of encapsulated shoot tips into plantlets also occurred when calcium alginate beads were directly sown in autoclaved soilrite moistened with 1/4-MS salts. Encapsulation of vegetative propagules in calcium alginate beads can be used as an alternative to synthetic seeds derived from somatic embryos.     

Differentiation of bone marrow-derived mesenchymal stem cells into hepatocyte-like cells in an alginate scaffold

Objectives: Alginate scaffolds are the most frequently investigated biomaterials in tissue engineering. Tissue engineering techniques that generate liver tissue have become important for treatment of a number of liver diseases and recent studies indicate that bone marrow‐derived stem cells (BMSCs) can differentiate into hepatocyte‐like cells. The goal of the study described here, was to examine in vitro hepatic differentiation potential of BMSCs cultured in an alginate scaffold. Materials and methods: To investigate the potential of BMSCs to differentiate into hepatocyte‐like cells, we cultured BMSCs in alginate scaffolds in the presence of specific growth factors including hepatocyte growth factor, epidermal growth factor and fibroblast growth factor‐4. Results: We can demonstrate that alginate scaffolds are compatible for growth of BMSCs and when cultured in alginate scaffolds for several days they display several liver‐specific markers and functions. Specifically, they expressed genes encoding alpha‐foetoprotein, albumin (ALB), connexin 32 and CYP7A1. In addition, these BMSCs produced both ALB and urea, expressed cytokeratin‐18 (CK‐18) and were capable of glycogen storage. Percentage of CK‐18 positive cells, a marker of hepatocytes, was 56.7%. Conclusions: Our three‐dimensional alginate scaffolds were highly biocompatible with BMSCs. Furthermore, culturing induced their differentiation into hepatocyte‐like cells. Therefore, BMSCs cultured in alginate scaffolds may be applicable for hepatic tissue engineering.     

Encapsulation of Immature Somatic Embryos of <i>Coffea arabica</i> L. for <i>in Vitro</i> Preservation

The present study aimed to develop a protocol for somatic embryogenesis and encapsulation of coffee embryos (Coffea arabica L.), for the conservation of genotypes with characteristics of commercial interest. Somatic embryos were induced from leaf explants in Murashige and Skoog medium (MS) supplemented with 1 mg · L⁻¹ of 2,4-dichlorophenoxiacetic acid (2,4-D) combined with 2 mg · L⁻¹ of benzyladenine (BA). Somatic embryos (SE) at the globular stage were encapsulated in a sodium alginate matrix; two treatments were tested: MS + 5 mg · L⁻¹ BA + 1 mg · L⁻¹ NAA + 3% (w/v) alginate, and MS + 7 mg · L⁻¹ BA + 5.7 mg · L⁻¹ indoleacetic acid (IAA) + 3% (w/v) alginate. Alginate was complexed with 100 mM calcium chloride (CaCl₂). Viability of the encapsulated SE was determined by staining with 0.01% fluorescein diacetate (FDA) after 0, 15, 30, and 45 days of storage at 4°C. Embryo viability was 100% in both treatments.     

Process optimization for microencapsulation of probiotic yeasts

Background

Microencapsulation is a technique which improves the survival and viability of probiotics. We demonstrate encapsulation of five potential probiotic yeasts with alginate and gum as encapsulation matrices to improve their gastrointestinal transit.

Methods

Gum extracted from various cereals viz. rice, oats, barley, finger millet and pearl millet along with alginate have been used to encapsulate five potential probiotic yeasts. Screening was carried out by measuring swelling index, encapsulation efficiency and nutritional value of microcapsules encapsulated with alginate and gum. The concentration of OBG, sodium alginate and inoculum dosage of probiotic yeasts was optimized using response surface methodology (RSM). Efficiency of alginate OBG microcapsules with or without coating materials viz. whey protein and chitosan also tested. The mucoadhesion ability and storage stability of alginate OBG microcapsules with coating materials were tested.

Results

Highest encapsulation efficiency of probiotic yeasts was noted using oats bran gum (OBG) microcapsules along with alginate in all the five probiotic yeasts. Notably whey protein coated microcapsules showed maximum GIT tolerance (95%) and mucoadhesion (90%) for L. starkeyi VIT-MN03. The minimum loss of viability was observed in L. starkeyi VITMN03 microcapsules on 60th day of storage.

Conclusions

This is the first report on optimization and survival of microencapsulated probiotic yeasts under simulated GIT conditions using natural gum and alginate as encapsulation matrices and whey protein as coating material.

     

Haemolymph glucose concentrations of juvenile rock lobsters, Jasus edwardsii, feeding on different carbohydrate diets

Postprandial changes in haemolymph glucose concentration ([Glc]H) were measured in 4-day-fasted juvenile intermoult spiny lobsters, Jasus edwardsii, provided with meals composed of glycogen, maltose, sucrose, glucose, or fructose in a gelatine base, or with gels of the algal glycans agar, alginate and carrageenan. Baseline [Glc]H was 0.61+/-0.02 mmol L(-1). After consumption of glycogen, maltose or sucrose, [Glc]H approximately doubled, peaked after 3 h and returned to baseline between 12 and 24 h. Glucose and fructose meals were followed by periods of sustained hyperglycaemia lasting more than 24 h (peaking at approximately 2.5 times baseline at 6 and 3 h respectively). Suggested explanations for augmented hyperglycaemic responses to glucose and fructose are: 1) these monosaccharides by-passed contact digestion and absorption in the R-cells of the digestive gland, directing them away from storage and toward transepithelial scavenging routes; or 2) glucose and fructose directly elicited release of crustacean hyperglycaemic hormone via a chemosensory reflex. Agar and alginate induced significant postprandial glycaemic responses, consistent with reports of carbohydrases in this species and indicating their potential for inclusion in artificial diets as both binders and energy sources. Carrageenan, a highly sulphated galactan, did not produce a glycaemic response. The measurement of glycaemic responses is a quick method of obtaining nutritional information on carbohydrates considered for inclusion in formulated diets prior to lengthy growth trials.     

海藻酸钙对骨质疏松症大鼠骨骼肌SDF-1含量和骨密度的影响

摘要 目的：探讨海藻酸钙对骨质疏松症大鼠骨骼肌基质细胞衍生因子-1（Stromal Cell-derived Factor-1,SDF-1）含量和骨密度的影响。方法：骨质疏松症大鼠（n=48）随机平分为三组-模型组、尼尔雌醇组与海藻酸钙组,在建模后1周后三组分别给予双蒸水、0.1 mg/100 g尼尔雌醇与37.5 mg/mL海藻酸钙/枸杞多糖凝胶微球水溶液灌胃治疗,1 次/d,检测大鼠骨骼肌SDF-1含量和骨密度变化情况。结果：（1）尼尔雌醇组与海藻酸钙组给药第4周与第8周的血清钙离子含量高于模型组（P<0.05）,磷离子含量低于模型组（P<0.05）,尼尔雌醇组与海藻酸钙组对比差异有统计学意义（P<0.05）;（2）尼尔雌醇组与海藻酸钙组给药第4周与第8周的骨骼肌SDF-1含量低于模型组（P<0.05）,海藻酸钙组低于尼尔雌醇组（P<0.05）;（3）尼尔雌醇组与海藻酸钙组给药第4周与第8周的腰椎和股骨骨密度高于模型组（P<0.05）,海藻酸钙组低于尼尔雌醇组（P<0.05）;（4）尼尔雌醇组与海藻酸钙组给药第4周与第8周的股骨最大载荷、最大应力高于模型组(P<0.05),海藻酸钙组高于尼尔雌醇组（P<0.05）;（5）海藻酸钙组造血细胞数量较多,骨皮质结构较完整,致密均匀粗壮,小梁数目明显增多,骨髓腔变小。结论：海藻酸钙在骨质疏松症大鼠的应用能抑制骨骼肌SDF-1的释放,有助于提高骨密度,改善骨生物力学指标,提高血清钙离子含量,降低磷离子含量。     

Luminescent yeast cells entrapped in hydrogels for estrogenic endocrine disrupting chemical biodetection

In the construction of luminescent yeast cell based fibre-optic biosensors, we demonstrate a novel approach for estrogenic endocrine disrupting chemical (EDC) biodetection by entrapping genetically modified Saccharomyces cerevisiae cells, containing the estrogen receptor alpha-mediated expression of the luc reporter gene, in hydrogel matrices based on calcium alginate or PVA. In order to insure a significant signal, an optimal immobilization ratio of 1:2 alginate 3% (w/v): 5 × 10⁶ [cells/ml], respectively, was used with the highest 17-β-estradiol (β-E2) induction factor after 2.5 h of incubation with 10 [nM] β-E2. It was shown that biocompatible alginate beads, 4.27–4.55 × 10⁵ [CFU/bead], which were characterized by a detection limit of 0.08 [μg l⁻¹] and an EC50 of 0.64 [μg l⁻¹] for β-E2, retained their viability for luminescence measurements after 1 month of storage at −80 °C slow freeze condition, and thus repeated cell cultivations were not required. The assay reproducibility for each tested EDC, represented by the coefficients of variation (CV), ranged from 4.35 to 18.47%. An alternative immobilization method, based on a room temperature partial drying of polyvinyl alcohol (PVA) solution (LentiKat^® Liquid) and cell suspension mix, was investigated with only a slightly lower detection limit for β-E2 than that reported with alginate beads. Alginate yeast based hydrogels may also be applicable to the analysis of environmental water samples since the trend of detected estrogenic activities with alginate beads roughly correlated with LC–MS–MS analytical results.     

Co-entrapment of Trichoderma harzianum and Glomus sp. within alginate beads: impact on the arbuscular mycorrhizal fungi life cycle

Aims: This study was performed to explore the compatibility and applicability of plant beneficial micro‐organisms (i.e. Trichoderma harzianum MUCL 29707 and Glomus sp. MUCL 41833) co‐entrapped in alginate beads. Methods and Results: Spores of Glomus sp. and conidia of T. harzianum were immobilized in alginate beads and the impacts of the saprotrophic fungi on the presymbiotic and symbiotic phase of the arbuscular mycorrhizal (AM) fungi evaluated under strict in vitro culture conditions. Our results demonstrated the capacity of both micro‐organisms in combination to regrowth outside the calcium alginate coating. The presence of T. harzianum did not hinder the AM fungal development but rather stimulated its spore production and fitness. Conclusions: The combination of T. harzianum MUCL 29707 with Glomus sp. MUCL 41833 in alginate beads may represent a reliable alternative inoculum formulation for application in sustainable agriculture. Significance and Impact of the Study: The entrapment in the alginate beads of two fungi (i.e. a saprotroph and a symbiont) having beneficial effects on plants represents a promising formulation for the development of inoculants adapted to field application.     

Critical review of activated sludge modeling: State of process knowledge,modeling concepts,and limitations

This work critically reviews modeling concepts for standard activated sludge wastewater treatment processes (e.g., hydrolysis, growth and decay of organisms, etc.) for some of the most commonly used models. Based on a short overview on the theoretical biochemistry knowledge this review should help model users to better understand (i) the model concepts used; (ii) the differences between models, and (iii) the limits of the models. The seven analyzed models are: (1) ASM1; (2) ASM2d; (3) ASM3; (4) ASM3 + BioP; (5) ASM2d + TUD; (6) Barker & Dold model; and (7) UCTPHO+. Nine standard processes are distinguished and discussed in the present work: hydrolysis; fermentation; ordinary heterotrophic organisms (OHO) growth; autotrophic nitrifying organisms (ANO) growth; OHO & ANO decay; poly‐hydroxyalkanoates (PHA) storage; polyphosphate (polyP) storage; phosphorus accumulating organisms PAO) growth; and PAO decay. For a structured comparison, a new schematic representation of these processes is proposed. Each process is represented as a reaction with consumed components on the left of the figure and produced components on the right. Standardized icons, based on shapes and color codes, enable the representation of the stoichiometric modeling concepts and kinetics. This representation allows highlighting the conceptual differences of the models, and the level of simplification between the concepts and the theoretical knowledge. The model selection depending on their theoretical limitations and the main research needs to increase the model quality are finally discussed. Biotechnol. Bioeng. 2013; 110: 24–46. © 2012 Wiley Periodicals, Inc.     

Penicillin G production by immobilized fungal vesicles

Summary The activity of penicillin G production was compared in a well defined medium by native vesicles as well as by calcium alginate gel immobilized vesicles isolated from the protoplasts of Penicillium chrysogenum PQ-96. The activity yield of the immobilized vesicles was 44% in comparison with native vesicles. After 60 h of storage at 4° C the native vesicles showed a rapid decrease in penicillin G production. The storage stability of these vesicles was improved after entrapment inside the calcium alginate gel. After 240 h of storage 1 mg of immobilized vesicular protein catalyzed the production of about 140 nmoles of penicillin G in 1 h.     

Manipulation of the acetylation degree of Azotobacter vinelandii alginate by supplementing the culture medium with 3-(N-morpholino)-propane-sulfonic acid

AIMS: The aim of this study was to characterize the influence of 3-(N-morpholino)-propane-sulfonic acid (MOPS) on alginate production by Azotobacter vinelandii and its chemical composition (particularly its acetylation degree), as well as on the rheological behaviour of alginate-reconstituted solutions. METHODS AND RESULTS: Cultures were grown in 500-ml flasks containing 90 ml of medium supplemented with MOPS in concentrations ranging from 0 to 13.6 mmol l(-1). The acetylation degree of the alginate was significantly influenced by the MOPS concentration, obtaining an alginate with an acetylation degree of 1.4% when 13.6 mmol l(-1) of MOPS was added to the medium. This value was twice as high as that obtained when no MOPS was used. The higher acetylation of the polymer resulted in higher viscosity of alginate solutions, having a more pronounced pseudoplastic behaviour. CONCLUSIONS: MOPS added to the culture medium determines the acetyl content of the alginate and thus, the physico-chemical properties of the polymer. SIGNIFICANCE AND IMPACT OF THE STUDY: These changes in the functional properties of the polymer can be very valuable in specific applications of alginate in the food and pharmaceutical fields.     

The preservation of micro-organisms in biological specimens stored at-70 degrees C

The preservation of micro-organisms that may be found on the skin was studied by storage in liquid media at--70 degrees C. In the first part of the study the performance of 12 varieties of suspending media was evaluated with pure cultures of 17 species of micro-organisms maintained in the laboratory. After storage for 1 year the best medium (Oxoid Nutrient Broth with 15% glycerol) showed a mean survival for all organisms studied of 83.8%, with no significant differences between organisms. Even the worst medium (distilled water) permitted greater than 40% survival at 1 year. No changes in the characteristics of these micro-organisms were detected after 6 months storage in glycerol broth. In the second part of the study nose swabs were suspended in one representative medium (Bacto Nutrient Broth containing 7% glycerol). The mean percentage survival of staphylococci in these suspensions after 1 year's storage at - 70 degrees C was 75.4%. These results indicate that coagulase-negative coagulase-negative staphylococci in samples of skin flora may be stored under these conditions for long periods, greatly reducing the work-load in epidemiological studies of infection.     

The preservation of micro-organisms in biological specimens stored at – 70°C

The preservation of micro-organisms that may be found on the skin was studied by storage in liquid media at -70°C. In the first part of the study the performance of 12 varieties of suspending media was evaluated with pure cultures of 17 species of micro-organisms maintained in the laboratory. After storage for 1 year the best medium (Oxoid Nutrient Broth with 15% glycerol) showed a mean survival for all organisms studied of 83.8%, with no significant differences between organisms. Even the worst medium (distilled water) permitted greater than 40% survival at 1 year. No changes in the characteristics of these micro-organisms were detected after 6 months storage in glycerol broth. In the second part of the study nose swabs were suspended in one representative medium (Bacto Nutrient Broth containing 7% glycerol). The mean percentage survival of staphylococci in these suspensions after 1 year's storage at -70°C was 75.4%. These results indicate that coagulase-negative staphylococci in samples of skin flora may be stored under these conditions for long periods, greatly reducing the work-load in epidemiological studies of infection.     

Distribution of alginate gene sequences in the Pseudomonas rRNA homology group I-Azomonas-Azotobacter lineage of superfamily B procaryotes.

Chromosomal DNA from group I Pseudomonas species, Azotobacter vinelandii, Azomonas macrocytogens, Xanthomonas campestris, Serpens flexibilis, and three enteric bacteria was screened for sequences homologous to four Pseudomonas aeruginosa alginate (alg) genes (algA, pmm, algD, and algR1). All the group I Pseudomonas species tested (including alginate producers and nonproducers) contained sequences homologous to all the P. aeruginosa alg genes used as probes, with the exception of P. stutzeri, which lacked algD. Azotobacter vinelandii also contained sequences homologous to all the alg gene probes tested, while Azomonas macrocytogenes DNA showed homology to all but algD. X. campestris contained sequences homologous to pmm and algR1 but not to algA or algD. The helical bacterium S. flexibilis showed homology to the algR1 gene, suggesting that an environmentally responsive regulatory gene similar to algR1 exists in S. flexibilis. Escherichia coli showed homology to the algD and algR1 genes, while Salmonella typhimurium and Klebsiella pneumoniae failed to show homology with any of the P. aeruginosa alg genes. Since all the organisms tested are superfamily B procaryotes, these results suggest that within superfamily B, the alginate genes are distributed throughout the Pseudomonas group I-Azotobacter-Azomonas lineage, while only some alg genes have been retained in the Pseudomonas group V (Xanthomonas) and enteric lineages.     

Distribution of alginate gene sequences in the Pseudomonas rRNA homology group I-Azomonas-Azotobacter lineage of superfamily B procaryotes.

Chromosomal DNA from group I Pseudomonas species, Azotobacter vinelandii, Azomonas macrocytogens, Xanthomonas campestris, Serpens flexibilis, and three enteric bacteria was screened for sequences homologous to four Pseudomonas aeruginosa alginate (alg) genes (algA, pmm, algD, and algR1). All the group I Pseudomonas species tested (including alginate producers and nonproducers) contained sequences homologous to all the P. aeruginosa alg genes used as probes, with the exception of P. stutzeri, which lacked algD. Azotobacter vinelandii also contained sequences homologous to all the alg gene probes tested, while Azomonas macrocytogenes DNA showed homology to all but algD. X. campestris contained sequences homologous to pmm and algR1 but not to algA or algD. The helical bacterium S. flexibilis showed homology to the algR1 gene, suggesting that an environmentally responsive regulatory gene similar to algR1 exists in S. flexibilis. Escherichia coli showed homology to the algD and algR1 genes, while Salmonella typhimurium and Klebsiella pneumoniae failed to show homology with any of the P. aeruginosa alg genes. Since all the organisms tested are superfamily B procaryotes, these results suggest that within superfamily B, the alginate genes are distributed throughout the Pseudomonas group I-Azotobacter-Azomonas lineage, while only some alg genes have been retained in the Pseudomonas group V (Xanthomonas) and enteric lineages.