Rheological Characterization and Drug Release Studies of Gum Exudates of Terminalia catappa Linn

The present study was undertaken to evaluate the gum exudates of Terminalia catappa Linn. (TC gum) as a release retarding excipient in oral controlled drug delivery system. The rheological properties of TC gum were studied and different formulation techniques were used to evaluate the comparative drug release characteristics. The viscosity was found to be dependent on concentration and pH. Temperature up to 60°C did not show significant effect on viscosity. The rheological kinetics evaluated by power law, revealed the shear thinning behavior of the TC gum dispersion in water. Matrix tablets of TC gum were prepared with the model drug dextromethorphan hydrobromide (DH) by direct compression, wet granulation and solid dispersion techniques. The dissolution profiles of the matrix tablets were compared with the pure drug containing capsules using the USP Basket apparatus with 500 ml phosphate buffer of pH 6.8 as a dissolution medium. The drug release from the compressed tablets containing TC gum was comparatively sustained than pure drug containing capsules. Even though all the formulation techniques showed reduction of dissolution rate, aqueous wet granulation showed the maximum sustained release of more than 8 h. The release kinetics estimated by the power law revealed that the drug release mechanism involved in the dextromethorphan matrix is anomalous transport as indicated by the release exponent n values. Thus the study confirmed that the TC gum might be used in the controlled drug delivery system as a release-retarding polymer.     

Characterization of Carrageenan Extracted from <Emphasis Type="Italic">Hypnea bryoides</Emphasis> in Oman

Carrageenophyte red seaweed from Oman, Hypnea bryoides, extracted using three different processes: an aqueous, a mild alkaline, and a more vigorous alkaline extraction was investigated. The resulting extract precipitated by alcohol was subject to chemical and rheological measurements. The total carbohydrate [ranged from 36.78 to 41.65 g/100 g], and ash [39.04 to 43.11 g/100 g] were the most abundant components in H. bryoides and contrary to the two, lipid content was found at a minimum [ranging from 2.95 to 3.38 g/100 g]. Alkali treatment with NaOH allowed complete conversion of kappa (κ) carrageenan form as detected by FTIR analysis. Total yield by alkali treatments gave higher yields (33%) compared with aqueous treatments (12%). However, subsequent aqueous treatment produced mixed carrageenan (μ and κ) with higher molecular weight compared with the alkali treatments which produced single carrageenan form (κ) with molecular weight of 4.1 × 10⁵ Da. The effects of thermal history on gel–sol and sol–gel transition were investigated by differential scanning calorimeter (DSC) and rheology on a pure sample and 1.5% κ-carrageenan mixture added with 30 mM KCl. Transition temperatures from DSC and rheology showed comparable results and were in good agreement with those previously reported.     

Protoplasts from <Emphasis Type="Italic">Agrobacterium rhizogenes</Emphasis>-transformed cell line of <Emphasis Type="Italic">Medicago sativa</Emphasis> L. regenerated to hairy roots

Summary Protoplasts were isolated from Agrobacterium rhizogenes A4-transformed cell line of Medicago sativa L. The highest yield of protoplasts (4.2×10⁶ per g fresh weight) was obtained from 12-d-old calluses after being subeultured on fresh medium. The viability of protoplasts reached over 80%. Protoplasts were induced to undergo sustained divisions when cultured in Durand et al. (DPD) medium supplemented with 2 mgl⁻¹ (9.05 μM) 2,4-dichlorophenoxyacetic acid, 0,2mgl⁻¹ (0.93 μM) kinetin, 0.3 M mannitol, 2% (w/v) sucrose, and 500 mgl⁻¹ casein hydrolyzate at a plating density of 1.0×10⁵ per ml. An agarose-beads culture method was appropriate for protoplast division of transformed alfalfa. The division frequency was about 30%. Numerous hairy roots were induced from protocalluses on Murashige and Skoog medium without growth regulators. Paper electrophoresis revealed that all of the regenerated hairy roots tested synthesized the corresponding opines. This protoplast culture system would be valuable for further somatic hybridization in forage legumes.     

An energy budget for Simocephalus vetulus (O. F. Muller) (Crustacea : Cladocera)

We discuss the energetics of a cladoceran, Simocephalus vetulus at different temperatures (8.0 ± 1.0, 15.0 ± 1.0, 21.0 ± 1.0 and 28.0 ± 1.0 °C) and food (Chlamydomonas sp.) concentrations (25 × 10³, 50 × 10³, 75 × 10³ and 100 × 10³ cells ml⁻¹). Increase in temperature accelerated ingestion and, to some extent, oxygen consumption. The study revealed a high reproduction efficiency in S. vetulus. Net growth efficiency (ECI) was higher (13.17–41.18%) in pre-adults than in adults (2.71–8.40%). The assimilated energy (A) increased with increasing food concentration at all temperatures. Assimilation efficiency (AD) decreased with increasing food concentrations. The energy used for growth (P) was nearly constant at all food levels because the egested energy increased and assimilation efficiency decreased as food concentration increased.     

Production,purification and partial characterisation of a novel laccase from the white-rot fungus <Emphasis Type="Italic">Panus tigrinus</Emphasis> CBS 577.79

Extracellular laccase from Panus tigrinus CBS 577.79 was produced in a bubble-column reactor using glucose-containing medium supplemented with 2,5-xylidine under conditions of nitrogen sufficiency. The main laccase isoenzyme was purified to apparent homogeneity by ultra-filtration, anion-exchange chromatography and gel filtration that led to a purified enzyme with a specific activity of 317 IU (mg protein)⁻¹ and a final yield of 66%. Laccase was found to be a monomeric protein with a molecular mass of 69.1 kDa, pI of 3.15 and 6.9% N-glycosylation of the high mannose type. Temperature and pH optima were 55°C and 3.75 (2,6-dimethoxyphenol as substrate). At 50 and 60°C, the enzyme half-lives were 281 and 25 min, respectively. The P. tigrinus laccase oxidized a wide range of both naturally occurring and synthetic aromatic compounds: the highest catalytic efficiencies were for 2,2′-azinobis-(3-ethylbenzthiazoline-6-sulfonic) acid and 2,6-dimethoxyphenol (5.99 × 10⁶ and 3.07 × 10⁶ M⁻¹ s⁻¹, respectively). Catalytic rate constants for typical N–OH redox mediators, such as 1-hydroxybenzotriazole (2.6 s⁻¹), violuric acid (8.4 s⁻¹) and 2,2,6,6-tetramethylpiperidin-N-oxide radical (7.8 s⁻¹), were found to be higher than those reported for other high redox potential fungal laccases.     

High efficiency <Emphasis Type="Italic">In vitro</Emphasis> plant regeneration from cotyledon explants of <Emphasis Type="Italic">Incarvillea sinensis</Emphasis>

Summary A simple and effective procedure has been developed for plantlet regeneration from cotyledon-derived callus of the medicinally important herb and ornamental species, Incarvillea sinensis. An average of 18.4 adventitious shoots per explant were obtained from 100% cotyledon explants cultured on half-strength Murashige and Skoog (MS) medium containing 1.0 mg l⁻¹ 6-benzylaminopurine for 3 wk, followed by another 4 wk on hormone-free 1/2×MS medium. The cotyledon explants continued to expand and regenerate new shoots upon repeated subculturing onto fresh medium. Most regenerated shoots (66.9%) were rooted on 1/4×MS mediumcontaining 1.0 mg l⁻¹ indole-3-acetic acid, with an average of about 3.8 roots per shoot. Regenerated plants with well developed shoots and roots were successfully acclimatized in soil and were normal phenotypically.     

Mucoadhesive, thermosensitive, prolonged-release vaginal gel for clotrimazole: β-cyclodextrin complex

The purpose of this study was to achieve a better therapeutic efficacy and patient compliance in the treatment for vaginitis. Clotrimazole (1%) has been formulated in a vaginal gel using the thermosensitive polymer Pluronic F127 (20%) together with mucoadhesive polymers such as Carbopol 934 and hydroxypropylmethylcellulose (0.2% for both). To increase its aqueous solubility., clotrimazole was incorporated as its inclusion complex with 1∶1 molar ratio with β-cyclodextrin. The inclusion complex was thoroughly characterized using various techniques, including ¹H NMR spectroscopy, FT IR spectrophotometry, differential scanning calorimetry, scanning electron microscopy, phase solubility studies, and determination of stability constant (k_1∶1). The gelation temperature and rheological behavior of different formulations at varying temperatures were measured. In vitro release profiles of the gels were determined in pH 5.5 citrate buffer. It was observed that complexation with cyclodextrin slowed down the release of clotrimazole considerably. Carbopol 934, on the other hand, was found to interact with β-cyclodextrin, inducing precipitation. As far as rheological properties are concerned, thermosensitive in situ gelling was obtained with formulations containing drug: cyclodextrin complex rather than with free drug. Thus, the optimum formulation for a controlled-release thermosensitive and mucoadhesive vaginal gel was determined to be clotrimazole: β-cyclodextrin 1% with 0.2% hydroxypropylmethylcellulose in Pluronic F127 gel (20%) providing continuous and prolonged release of active material above MIC values.     

Elicitation of guggulsterone production in cell cultures of <Emphasis Type="Italic">Commiphora wightii</Emphasis> by plant gums

Plant gum as an elicitor for guggulsterone production in cell cultures of Commiphora wightii is reported for the first time. Guggulsterone production increased 2.4 fold in the cell cultures by gum Arabic (100 mg l⁻¹), while mesquite gum elicited 2 fold. The cells treated with gum Arabic at 7th and 9th day accumulated enhanced guggulsterones within 24 h, which increased further up to 48 h and then declined. The cells treated at 9th day accumulated higher amount (218 μg l⁻¹) of guggulsterones after 48 h of elicitation as compared to cells treated at 7th day (164 μg l⁻¹). The optimized elicitation conditions were used in vessels of varying capacity where maximum yield of 285 μg l⁻¹ of guggulsterones was recorded in 3 l shake flasks. These experiments enabled highest guggulsterones yield in a short duration of 11 days in cell cultures of C. wightii.     

Combined effects of the entomopathogenic fungus, <Emphasis Type="Italic">Paecilomyces fumosoroseus</Emphasis> Apopka-97, and the generalist predator, <Emphasis Type="Italic">Dicyphus hesperus</Emphasis>, on whitefly populations

The effects of intraguild interactions between Dicyphus hesperus Knight (Hemiptera: Miridae) and Paecilomyces fumosoroseus Apopka-97 (PFR-97^TM) (Wize) Brown and Smith (Ascomycota: Hypocreales) on Trialeurodes vaporariorum (Westwood) (Hemiptera: Aleyrodidae) populations were investigated in tomato greenhouse microcosms. Conditions were established in which interference or synergy would most likely occur; namely, a high number of available whiteflies were combined with large numbers of both D. hesperus and PFR-97^TM. We measured live whitefly density in a factorial repeated measures experiment where plants were provided or withheld releases of D. hesperus and/or applications of PFR-97^TM for 6 weeks. Releases of D. hesperus were made at a rate of 10 adults/plant during the first and third week and PFR-97^TM suspensions were applied with a backpack sprayer at a rate of 18 × 10⁷, 1.3 × 10⁷ and 1.2 × 10⁷ viable blastospores/ml during the first, third and fourth week, respectively. Results revealed a non-significant interaction effect between D. hesperus and PFR-97^TM, indicating that their actions were independent. Individual whitefly reductions of 48% and 35% were achieved by PFR-97^TM and D. hesperus, respectively. Collectively, the natural enemies reduced whitefly densities by 62% relative to the controls. The density of D. hesperus adults was unaffected by multiple applications of PFR-97^TM. These results suggest that the combination of generalist entomopathogenic fungi and generalist predators has the potential to cause increased pest mortality despite evidence of minimal interference.     

Batch fermentation with entrapped growing cells ofLactobacillus casei

Summary Growing cells ofLactobacillus casei were entrapped in-carrageenan/locust bean gum (LBG) (2:1 or 2.75%:0.25% w/w respectively) mixed gel beads (two ranges of diameter: 0.5–1.0 and 1.0–2.0 mm) to fermentLactobacillus Selection (LBS) medium and produce biomass. The results showed significant influence of initial cell loading of the beads and bead size on the fermentation rate. The highest cell release rates were obtained with 2.75%:0.25%-carrageenan/LBG small diameter gel beads. However, 17 h fermentation of LBS medium with immobilized cells resulted in substantial softening of the gel matrix, prohibiting reuse of immobilized biocatalysts as inoculum in subsequent batch fermentation. A dynamic shear rheological study showed that the gel weakness was related to chemical interactions with the medium. Results indicated that part of the matrix-stabilizing K⁺ ions diffused back to the medium. Stabilization of the gel was obtained by adding potassium ions to the LBS medium;L. casei growth was not altered by this supplementation. Fermentation of LBS medium supplemented with KCl byL. casei showed higher cell counts in the broth medium with immobilized cells than with free cells, reaching 10¹⁰ cells/ml after about 10 h with entrapped cells in 0.5–1.0 mm diameter beads and 17 h with free cells. Counts in the gel beads after fermentation were higher than 10¹¹ cells/ml and bead integrity was maintained throughout fermentation.     

Gellan gum gel as entrapment matrix for high temperature fermentation processes: A rheological study

High temperature rheological measurements (60–80°C) were carried out on gellan gum gel. The experiments were conducted in order to optimize the behavior of the gel as an entrapment matrix for fermentation processes of dairy fluids using highly thermophilic microorganisms. At 60°C, no significant decrease of rheological parameters of the gel was observed for 15 days, as compared to 23°C. At 80°C, the gel weakened abruptly. The gel was extremely sensitive to calcium loss (optimum concentration 0.1% CaCl₂) due to chelating agents. Gellan gum gel could be an efficient entrapment matrix for fermentation processes at temperature up to 60°C in dairy fluids.     

Rheological properties of <Emphasis Type="Italic">Chlorella pyrenoidosa</Emphasis> culture grown heterotrophically in a fermentor

Rheological properties of Chlorella pyrenoidosa culture grown heterotrophically in a 14 L fermentor were investigated. It was found that the fluid viscosity was rather low and remained almost unchanged during the cultivation, implying that no (or very few) viscous substances were excreted into the medium. Investigation of the condensed suspension of C. pyrenoidosa showed that for biomass concentration under 150 g.L⁻¹, the suspension of C. pyrenoidosa exhibited Newtonian behavior, and the fluid viscosity was rather low (about 40 mPa·s) and increased very slowly with the increase in cell concentration. With further increase in biomass concentration however, the fluid rheological behavior changed to non-Newtonian, and the fluid viscosity increased rapidly with the increase in cell concentration. From the viewpoint of rheology, C. pyrenoidosa is an excellent organism for high-cell-density culture, and there will be no rheological problems at cell densities under 150 g.L⁻¹.     

Effect of Concentration and Temperature on Flow Properties of Alyssum homolocarpum Seed Gum Solutions: Assessment of Time Dependency and Thixotropy

Effects of different shear rates (14, 25, and 50 s⁻¹), gum concentrations (3%, 3.5%, and 4%), and temperatures (5–65 °C) on flow properties of Alyssum homolocarpum seed gum solutions were investigated using a rotational viscometer. The experimental data were fitted with three time-dependent rheological models, namely second-order structural kinetic model, Weltman model, and first-order stress decay model with a non-zero stress value. The rate constant and extent of viscosity strongly depended on the shear rate, gum concentration, and temperature. It was found that A. homolocarpum seed gum samples exhibited shear thinning and thixotropic behavior for all concentrations and temperatures. The amount of structural breakdown decreased with shear rate, but it did not have a general trend with concentration and temperature. The extent of thixotropy increased with increasing gum concentration and decreased with increasing temperature and shear rate. In this work, the decay rate constant generally increased with increasing shear rate; however, it did not have any trend with concentration and temperature.     

Evaluation of <Emphasis Type="BoldItalic">Sterculia foetida</Emphasis> Gum as Controlled Release Excipient

The purpose of the research was to evaluate Sterculia foetida gum as a hydrophilic matrix polymer for controlled release preparation. For evaluation as a matrix polymer; characterization of Sterculia foetida gum was done. Viscosity, pH, scanning electronmicrographs were determined. Different formulation aspects considered were: gum concentration (10–40%), particle size (75–420 μm) and type of fillers and those for dissolution studies; pH, and stirring speed were considered. Tablets prepared with Sterculia foetida gum were compared with tablets prepared with Hydroxymethylcellulose K15M. The release rate profiles were evaluated through different kinetic equations: zero-order, first-order, Higuchi, Hixon-Crowell and Korsemeyer and Peppas models. The scanning electronmicrographs showed that the gum particles were somewhat triangular. The viscosity of 1% solution was found to be 950 centipoise and pH was in range of 4–5. Suitable matrix release profile could be obtained at 40% gum concentration. Higher sustained release profiles were obtained for Sterculia foetida gum particles in size range of 76–125 μm. Notable influences were obtained for type of fillers. Significant differences were also observed with rotational speed and dissolution media pH. The in vitro release profiles indicated that tablets prepared from Sterculia foetida gum had higher retarding capacity than tablets prepared with Hydroxymethylcellulose K15M prepared tablets. The differential scanning calorimetry results indicated that there are no interactions of Sterculia foetida gum with diltiazem hydrochloride. It was observed that release of the drug followed through surface erosion and anomalous diffusion. Thus, it could be concluded that Sterculia foetida gum could be used a controlled release matrix polymer.     

Microalgal growth enhancement by levoglucosan isolated from the green seaweed <Emphasis Type="Italic">Monostroma nitidum</Emphasis>

Microalgal growth was enhanced by the addition of levoglucosan to the culture medium. The growth-enhancing compound levoglucosan was isolated from the green seaweed Monostroma nitidum using water extraction, molecular fractionation, DEAE-cellulose column chromatography, and high-performance liquid chromatography. Yield of the compound from seaweed powder was 5 × 10⁻³% (w/w). At 10 mM concentration, levoglucosan enhanced cell growth and the specific growth rate of all feed microalgal species tested (Chaetoceros gracilis, Chlorella ellipsoidea, Dunaliella salina, Isochrysis galbana, Nannochloris oculata, Navicula incerta, Pavlova lutheri, Tetraselmis suecica) in most culture media by approximately 150%. Cellular fatty acid profiles and cell size differed marginally between cultures with and without levoglucosan.     

Plant regeneration from leaf protoplasts of <Emphasis Type="Italic">Solanum virginianum</Emphasis> L. (<Emphasis Type="Italic">Solanaceae</Emphasis>)

Leaves of Solanum virginianum plants were used for protoplast isolation. To support cell wall formation and cell division, protoplasts were cultured in thin alginate layers floated in liquid medium. When protoplasts were plated at a density of 1.0 × 10⁶/ml in Kao and Michyaluk (KMp8) medium supplemented with 0.5 mg/l zeatin, 1.0 mg/l 2,4-dichlorophenoxyacetic acid, and 1.0 mg/l α-naphthaleneacetic acid, 42.3% of the dividing cells developed microcalli in 3–4 weeks. Shoot formation via organogenesis of protoplast-derived calli was achieved for 28% of calli transferred to solidified KMp8 medium supplemented with 2.0 g/l zeatin and 0.1 mg/l 3-indol acetic acid in about 2 weeks. Further shoot development was observed in Murashige and Skoog (MS) medium without growth regulators and roots were induced after transfer to MS medium containing 1.0 mg/l 3-indol butyric acid. Regenerated plants have normal morphology.     

Physicochemical and Rheological Characterization of Gum Tragacanth Exudates from Six Species of Iranian <Emphasis Type="Italic">Astragalus</Emphasis>

The flow behavior of six species of Iranian gum tragacanth dispersions was investigated at different temperatures and ionic strengths, within a concentration range (0.05–1.5% w/w) using a controlled shear rate rheometer. The steady shear measurements showed that all of the gum dispersions had shear-thinning natures. The power law model was used to describe the rheological properties of dispersions and Arrhenius model was used to evaluate the temperature effect. Composition analysis, surface tension measurement, particle size analysis, and color measurement of all the species were also carried out. The results indicated that the six species of gum tragacanth being studied in this paper exhibited significantly different physicochemical properties. Therefore, various species of gum tragacanth can be used instead of different hydrocolloids in a wide range of applications.     

Measurement of particle diameter of <Emphasis Type="Italic">Lactobacillus acidophilus</Emphasis> microcapsule by spray drying and analysis on its microstructure

Lactobacillus acidophilus, as a probiotic, is widely used in many functional food products. Microencapsulation not only increases the survival rate of L. acidophilus during storage and extends the shelf-life of its products, but also optimal size microcapsule makes L. acidophilus have an excellent dispersability in final products. In this paper, L. acidophilus was microencapsulated using spray drying (inlet air temperature of 170°C; outlet air temperature of 85–90°C). The wall materials used in this study were β-cyclodextrin and acacia gum in the proportion of 9:1 (w/w), and microcapsules were prepared at four levels of wall materials (15, 20, 25 and 30% [w/v]) with a core material concentration of 6% (v/v). The microcapsule diameters were measured by Malvern’s Mastersizer-2000 particle size analyzer. The results showed that the particle diameters of microcapsule were mostly within 6.607 μm and 60.256 μm and varied with 2.884–120.226 μm (the standard smaller microcapsule designated as <350 μm). Through comparison of microcapsule size and uniformity with different concentration of wall materials, we concluded that the optimal concentration of wall material was 20% (w/v), which gave microcapsule with a relatively uniform size (averaging 22.153 μm), and the number of surviving encapsulated L. acidophilus was 1.50 × 10⁹ c.f.u./ml. After 8 weeks storage at 4°C, the live bacterial number was above 10⁷ c.f.u./ml, compared with unencapsulated L. acidophilus, 10⁴–10⁵ c.f.u./ml. Through the observation of scanning electron microscopy, we found that the shapes of microcapsule were round and oval, and L. acidophilus cells located in the centre of microcapsule.     

DNA-mediated transformation system in a bipolar basidiomycete, <Emphasis Type="Italic">Pholiota microspora</Emphasis> (<Emphasis Type="Italic">P. nameko</Emphasis>)

We cloned a gene encoding the succinate dehydrogenase iron-sulfur protein subunit (sip) from a bipolar mushroom, Pholiota microspora, and introduced a point mutation that confers carboxin resistance into this gene. Using this homologous selective marker and also a heterologous drug selective marker, the hygromycin B phosphotransferase gene (hph), we successfully constructed a DNA-mediated transformation system in P. microspora. Both these selection markers have high transformation efficiency: the efficiency of carboxin resistance transformation was about 88.8 transformants/μg pMBsip2 DNA using 5 × 10⁶ protoplasts in regeneration plates containing 1.0 μg/ml carboxin, and the efficiency of hygromycin B resistance transformation was about 122.4 transformants/μg pMBhph1 DNA using 5 × 10⁶ protoplasts in regeneration plates containing 150 μg/ml hygromycin B. Southern hybridization analysis showed that the introduced sequence (mutant sip or hph) was integrated into the chromosomal DNA in these transformants with a copy number of one or more.     

Seasonal abundance and distribution of <Emphasis Type="Italic">Vibrio</Emphasis> species in the treated effluent of wastewater treatment facilities in suburban and urban communities of Eastern Cape Province,South Africa

We assessed the seasonal abundance and distribution of Vibrio species as well as some selected environmental parameters in the treated effluents of two wastewater treatment plants (WWTP), one each located in a suburban and urban community of Eastern Cape Province, South Africa. Vibrio population density ranged from 2.1×10⁵ to 4.36×10⁴ CFU/ml in the suburban community and from 2.80×10⁵ to 1.80×10⁵ CFU/ml in the urban community. Vibrio species associated with 180 μ, 60 μ, and 20 μ plankton sizes were observed at densities of 0–136×10³ CFU/ml, 0–8.40×10² CFU/ml, and 0–6.80×10² CFU/ml, respectively at the suburban community’s WWTP. In the urban community, observed densities of culturable Vibrio were 0–2.80×10² CFU/ml (180 μ), 0–6.60×10² CFU/ml (60 μm), and 0–1.80× 10³ CFU/ml (20 μm). The abundance of free-living Vibrio species ranged from 0 to 1.0×10² and 1.0×10³ CFU/ml in the suburban and urban communities’ WWTPs, respectively. Molecular confirmation of the presumptive Vibrio isolates revealed the presence of V. fluvialis (41.38%), V. vulnificus (34.48%), and V. parahaemolyticus (24.14%) in the suburban community effluents. In the urban community molecular confirmation revealed that the same species were present at slightly different percentages, V. fluvialis (40%), V. vulnificus (36%), and V. parahaemolyticus (24%). There was no significant correlation between Vibrio abundance and season, either as free-living or plankton-associated entities, but Vibrio species abundance was positively correlated with temperature (r=0.565; p<0.01), salinity, and dissolved oxygen (p<0.05). Turbidity and pH showed significant seasonal variation (p<0.05) across the seasons in both locations. This study underscores the potential of WWTPs to be sources of Vibrio pathogens in the watershed of suburban and urban communities in South Africa.