Industrial wastewaters and dewatered sludge: rich nutrient source for production and formulation of biocontrol agent, <Emphasis Type="Italic">Trichoderma viride</Emphasis>

Axenic cultivation of biocontrol fungus Trichoderma viride was conducted on a synthetic medium and different wastewaters and wastewater sludges in shake flasks to search for a suitable raw material resulting in higher biocontrol activity. Soluble starch based synthetic medium, dewatered municipal sludge, cheese industry wastewater sludge, pre-treated and untreated pulp and paper industry wastewater and slaughter house wastewater (SHW) were tested for T. viride conidia and protease enzyme production. The maximum conidia production followed the order, soluble starch medium (>10⁹ c.f.u./mL), untreated pulp and paper industry wastewater (4.9 × 10⁷ c.f.u./mL) > cheese industry wastewater (1.88 × 10⁷ c.f.u./mL) ≈ SHW (1.63 × 10⁷ c.f.u./mL) > dewatered municipal sludge (3.5 × 10⁶ c.f.u./mL) > pre-treated pulp and paper industry wastewater (1.55 × 10⁶ c.f.u./mL). The protease activity of T. viride was particularly higher in slaughterhouse wastewater (2.14 IU/mL) and dewatered municipal sludge (1.94 IU/mL). The entomotoxicity of soluble starch based synthetic medium was lower (≈6090 SBU/μL) in contrast to other raw materials. The entomotoxicity inversely decreased with carbon to nitrogen ratio in the growth medium and the conidia concentration and protease activity also contributed to the entomotoxicity. The residual c.f.u./g formulation of T. viride conidia were up to approximately, 90% after 1 month at 4 ± 1 °C and about 70% after 6 months at 25 ± 1 °C. Thus, production of T. viride conidia would help in marketability of low cost biopesticide from the sludge and safe reduction of pollution load.     

Microbiological characterization of Civil cheese, a traditional Turkish cheese: microbiological quality, isolation and identification of its indigenous Lactobacilli

Summary The aim of this study was to investigate the microbiological quality and the indigenous lactic acid bacteria (Lactobacillus) of Civil cheese. In this study, for identification of lactic acid bacteria isolated from cheese samples, the Microbiology Identification System (MIS) was used. In the samples, average of total aerobic mesophilic bacteria, yeast and moulds, lactic acid bacteria and Staphylococcus aureus (Staph. aureus) were determined to be 3.0×10⁸, 2.0×10⁶, 1.4×10⁷, 4.10×10⁴ c.f.u./g, respectively. Coliform bacteria were lower than <10 c.f.u./g in 26.67 % of the samples. On the other hand, the coliform-positive samples showed an average of 4.2×10⁴ c.f.u./g. Staph. aureus was not detected in 33.33% of Civil cheese samples. Seventy-two strains of Lactobacillus isolated from Civil cheese were identified. Growth at 5, 10 and 37 °C, 2.5% and 6.5% NaCl and gas (CO₂) production from glucose of the isolated strains were also determined. Of the 72 isolates, 20 were identified as Lb. malefermentans, 18 as Lb. fermentum, 17 as Lb. parabuchneri, 10 as Lb. vaccinostercus, 2 as Lb. oris, 1 as Lb. bifermentans, 1 as Lb. delbruecki subsp. bulgaricus, 1 as Lb. cellobiosus, 1 as Lb. hilgardii, 1 as Lb. paracasei subsp. tolerans.     

Microbial population dynamics, especially stress tolerant Bacillus thuringiensis, in partially anaerobic rice field soils during post-harvest period of the Himalayan, island, brackish water and coastal habitats of India

Population ( × 10⁷ c.f.u./g dr. soil) of the aerobic (30.5–154.1) and anaerobic (5.9–91.4) heterotrophic, aerobic (24.0–56.0) and anaerobic (2.4–4.2) spore forming, Gram (-)ve (1.6–2.9), phosphate solubilizing (10–20), asymbiotic N₂-fixing (0.5–0.9), sulfur oxidizing (1.1–2.0), nitrifying (1.0–5.8) and denitrifying (12.1–18.7) bacteria; as well as, the actinomycetes (about 10⁴ c.f.u./g dr. soil) and fungi (about 10⁵ c.f.u./g dr. soil) were variable in the partially anaerobic, saline and drain out flooded rice soils during the post harvest period of the Himalayan, brackish water flooded, island and coastal habitats of India. The aerobic heterotrophic and spore forming bacteria were more than the anaerobic counterparts in the soils. Population (0.51–3.51 × 10⁶ c.f.u./g dr. soil) and crystal morphotype (spherical, bipyramidal and polymorphic) of the Bacillus thuringiensis (Bt) isolates of different soils were variable. Bt index was 0.002 at Mahe but 0.006 in other soils. The Bt isolates tolerated 5–12% NaCl. The osmolytes (mg/g dr. wt.) like the amino acids (0.38–99.45) and proline (0.38–0.80); and the antioxidative enzymes (units (U)/mg protein/min) viz. the catalase (0.17–5.59) and superoxide dismutase (0.35–74.46) were related with intrinsic osmotic stress tolerance of the Bt but they formed spores to overcome anoxic stress. Two Bt isolates were potent tolerant to both osmotic and anoxic stresses.     

Growth Profile and Hydrocarbonoclastic Potential of Microorganisms Isolated from Tarballs in the Bight of Bonny, Nigeria

Summary The growth profile and hydrocarbonoclastic potential of microorganisms isolated from tarballs harvested from Ibeno beach in the Bight of Bonny were examined to determine their role in the degradation of the aquatic pollutants (tarballs). The results of the analysis revealed that the mean heterotrophic bacterial count ranged from 3 (±0.01) × 10³ to 3.18 (±0.2) × 10⁵ c.f.u./g. The mycological count ranged from 1 (±0.3) × 10² to 2 (±0.4) × 10⁴ c.f.u./g while the mean count of biodegraders on tarball mineral salt medium (TMSM) ranged from 1 (±0.3) × 10² to 2 (±0.4) × 10⁴ c.f.u./g. The ability of the microbial isolates to utilize the tarballs as their sole source of carbon and energy was examined and noticed to vary in growth profiles between the isolates. Chromobacterium violaceum, Cladosporium resinae, Bacillus submarinus, Micrococcus varians, Pseudomonas aeruginosa, Candida marina and Saccharomyces estuari were the most efficient utilizers and biodegraders while Corynebacterium glutamicum, Nocardia marina, and Cryptococcus albidus exhibited moderate growth in TMSM. Vibrio parahaemolyticus and Escherichia coli were opportunistic inhabitants, as they could neither grow nor degrade the balls in TMSM. The results imply that the efficient biodegraders like Chromobacterium violaceum could extensively degrade the balls with time.     

Antivibrio compounds produced by <Emphasis Type="Italic">Pseudomonas</Emphasis> sp. W3: characterisation and assessment of their safety to shrimps

In order to explore compounds naturallly inhibitory to shrimp pathogenic vibrios, a culture filtrate of Pseudomonas sp. W3 at a pH of 2 was extracted with ethyl acetate (EtOAc) to produce 82.15 mg/l of a yellow–brown extract (EtOAc-W3) that had MIC values of 225-450 μg/ml against the growth of 18 shrimp pathogenic Vibrio harveyi strains. The MIC of EtOAc-W3 against the most pathogenic strain PSU 2015 was 450 μg/ml and this strain had the lowest LD₅₀ (50% lethal dose) to pacific white shrimp (Litopenaeus vannamei, PL 21). At this MIC value, EtOAc-W3 in artificial sea water (ASW) killed strain PSU 2015; however in natural sea water, only a partial growth inhibition was observed. The toxicity to pacific white shrimp and antivibrio activity of the EtOAc-W3 were investigated by conducting an experiment with 4 sets; native control (commercial ASW), EtOAc-W3 control (MIC/10, 45 μg/ml), challenge (inoculation 6.0 × 10⁶ c.f.u./ml PSU 2015) and treatment (6.0 × 10⁶ c.f.u./ml PSU 2015 + 45 μg/ml EtOAc-W3). The same experiment was repeated by increasing the dose of EtOAc-W3 to 90 μg/ml (MIC/5). Both concentrations of EtOAc-W3 tested had no toxicity to postlarval shrimps. A significant decrease in shrimp mortality was observed over a 72 h period as approximately 80% of the shrimps died in each challenge set but only 63 and 23% died in the presence of 45 and 90 μg/ml EtOAc-W3. The major component of EtOAc-W3 was supposed to be 2-heptyl-4-quinolone (HHQ) by FAB-MS and ¹H-NMR analyses of the purified fraction.     

Diversity of lactic acid bacteria and yeasts in Airag and Tarag,traditional fermented milk products of Mongolia

We used culture- and molecular-biology-based methods to investigate microbial diversity in the traditional Mongolian fermented milks “Airag” (fermented mare’s milk) and “Tarag” (fermented milk of cows, yaks, goats, or camels). By rRNA or functional gene sequencing, we identified 367 lactic acid bacteria (LAB) strains and 152 yeast strains isolated from 22 Airag and 31 Tarag samples. The total concentration of LAB in Airag (10^{7.78 ± 0.50} c.f.u. ml^–1; mean ± SD) was significantly lower (P < 0.01) than in Tarag (10^{8.35 ± 0.62} c.f.u. ml⁻¹), whereas the total concentration of yeasts in Airag (10^{7.41 ± 0.61} c.f.u. ml^-1) was significantly higher (P < 0.01) than in Tarag (10^{5.86 ± 1.29} c.f.u. ml^-1). Lactobacillus helveticus and Lactobacillus kefiranofaciens were isolated from Airag as the predominant LAB strains at levels of about 10⁷ c.f.u. ml⁻¹, whereas Lactobacillus delbrueckii subsp. bulgaricus, L. helveticus, and Streptococcus thermophilus were the predominant isolates from Tarag at about 10⁷ c.f.u. ml⁻¹. The lactose-fermenting Kluyveromyces marxianus was isolated predominantly from Airag as its major alcoholic fermentation component. Non-lactose-fermenting yeasts such as Saccharomyces cerevisiae, Issatchenkia orientalis, and Kazachstania unispora were the predominant isolates from Tarag, at about 10⁵ c.f.u. ml⁻¹. The apparent geographic differences in the L. kefiranofaciens and S. thermophilus contents of Tarag strongly suggested that differences among the animal species from which the milk was sourced, rather than geographic distances, were the most important factors influencing the diversity of the microbial composition of traditional fermented milks in Mongolia.     

Endophytic bacterial flora in the stem tissue of a tropical maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.) genotype: isolation,identification and enumeration

The genotypic diversity of indigenous bacterial endophytes within stem of tropical maize (Zea mays L.) was determined in field and greenhouse experiments. Strains were isolated from stem tissues of a tropical maize cultivar (PEHM-1) by trituration and surface disinfestation and their population dynamics was determined. Endophytes were found in most of the growing season at populations ranging from 1.36–6.12 × 10⁵ colony-forming units per gram fresh weight (c.f.u./gm fw) of stem. Analysis of these bacterial endophytes using Gas Chromatography—Fatty Acid Methyl Ester (GC-FAME) led to the identification of Bacillus pumilus, B. subtilis, Pseudomonas aeruginosa and P. fluorescens as the relatively more predominant group of bacterial species residing in maize stem. When the maize seedlings grown in a greenhouse were inoculated with these four isolates individually, their population densities decreased (1.6–3.1 × 10⁵ c.f.u./gm fw of stem) as compared to the field-grown maize (1.8–3.8 × 10⁵ c.f.u./gm fw of stem). The highest persistence, however, was recovered in the case of B. subtilis with a population density of 3.1 × 10⁵ c.f.u./gm fw of stem tissue on 28 days after emergence (DAE). This is the first report on population dynamics of bacterial endophytes from tropical maize and the results establish that symptomless populations of bacteria exist in the maize stem.     

Microbiological changes in mawè during natural fermentation

Lactic acid bacteria increased from 3.2 × 10⁶ and 1.6 × 10⁷ c.f.u./g (wet wt) to 2 × 10⁹ and 1.6 × 10⁹ c.f.u./g after 12 to 24 h of fermentation of home-produced mawè (a dough produced from dehulled maize) and commercial mawè, respectively. In commercial mawè, the yeast count increased from 1.3 × 10⁵ to 2.5 × 10⁷ c.f.u./g after 48 h of fermentation before decreasing, whereas in the home-produced mawè it increased from 2.5 × 10⁴ to 3.2 × 10⁷ c.f.u./g after 72 h of fermentation; the dominant yeasts were mainly Candida krusei, although C. kefyr, C. glabrata and Saccharomyces cerevisiae were also present. Enterobacteriaceae counts increased slightly during the initial stage ofthe fermentation, but decreased below the detection level after 24 to 48 h. Enterobacter cloacae was mostly found in commercial mawè and Escherichia coli mostly in homeproduced mawè.D.J. Hounhouigan and C.M. Nago are with the Université Nationale du Bénin, Faculté des Sciences Agronomiques, Département de Nutrition et de Sciences Alimentaires, BP 526, Cotonou, Benin; M.J.R. Nout and F.M. Rombouts are with the Agricultural University, Department of Food Science, Bomenweg 2, 6703 HD Wageningen, The Netherlands. J.H. Houben is with Utrecht University, Department of the Science of Foods of Animal Origin, Yalelaan 2, 3508 TD Utrecht, The Netherlands.     

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.     

Effect of curd-cooking temperatures on the microbiological quality ofayib,a traditional Ethiopian cottage cheese

Ayib was made following traditional methods of curd cooking at 40°, 50°, 60° and 70°C requiring 10 h, 7 h, 145 min and 55 min, respectively. During the natural fermentation of raw milk into sour milk (curd), the count of all groups of microorganisms increased by at least a 100-fold. When the curd was subsequently cooked at 40°C, counts of enterococci and members of Enterobacteriaceae decreased by a 100-fold and lactic acid bacteria and staphylococci by a 100-fold. At 50°C, the count of enterococci, members of Enterobacterlaceae and staphylococci was lower than could be detected (<10² c.f.u./g). Yeasts, moulds and lactic acid bacteria still had counts of about 10⁵ c.f.u./g. The total aerobic count decreased by a 100-fold but was still high (>10⁷ c.f.u./g). At 60°C, yeasts and moulds decreased by 1000-fold and at 70°C they decreased below detectable levels (<10² c.f.u./g). This latter temperature is thus recommended as it results in a less contaminated and saferayib.

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Résumé On a préparél'Ayib selon les méthodes traditionnelies de cuission du lait caillé à 40, 50, 60 et 70°C, ce qui a requis respectivement 10 h, 7 h, 145 et 55 min. Durant la fermentation naturelle du lait frais en lait caillé, le décompte de tous les groupes de microorganismes a crû d'au moins 100 fois. Quand le lait caillé était par la suite cuit à 40°C, le décompte des entérocoques et des membres des Enterobacteriaceae a décru de 100 fois et celui des bactéries lactiques et des staphylocoques de 100 fois. A 50°C, le décompte des entérocoques, membres des Enterobacteriaceae et des staphylocoques était inférieur au seuil de détection (<10² c.f.u./g). Les levures, moisissures et bactéries lactiques avalent toujours un décompte d'environ 10⁵ c.f.u./g. le décompte total aérobie décroissait de 100 fois mais restait néanmoins élevé (>10⁷ c.f.u./g.). A la température de cuisson du lait caillé de 60°C, les levures et les moisissures ont décru de 1000 fois, et à 70°C on décru au dessous du seuil de détection (<10² c.f.u./g). Cette dernière température est donc recommandable car elle résulte en unayib moins contaminé et plus sain.

     

Water-soluble granules containing <Emphasis Type="Italic">Bacillus megaterium</Emphasis> for biological control of rice sheath blight: formulation,bacterial viability and efficacy testing

Endospores of B. megaterium were formulated in granule formulations with sodium alginate, lactose and polyvinylpyrrolidone (PVP K-30) by the wet granulation technique. The granule formulation exhibited good physical characteristics, such as high-water solubility and optimal viscosity, that would be suitable for spray application. The bacteria remained viable in the dry granule formulation at 10⁹ c.f.u./g after 24 months storage at room temperature. Under laboratory conditions, aqueous solutions of the formulation showed high activity against mycelial growth of R. solani (99.64 ± 0.14% mycelial inhibition). High viability of the bacterial antagonist on leaf sheath and leaf blade at day 7 after spraying with the formulation was observed (approximately 10⁶ c.f.u./g of plant). Application of an equivalent number of un-formulated endospores resulted in much loss of the bacterial endospores even 1 day after application. In a small pilot field study, an aqueous solution of the formulation (3%w/v) applied by spraying at days 1, 5 and 10 after pathogen inoculation of the rice plants was more effective in suppressing rice sheath blight disease than one application of a fungicide (Iprodione) at day 1. Additionally, rice plants sprayed with the aqueous solution of the granule formulation had higher panicle and whole kernel weights than those of fungicide-treated and control (untreated) plants.     

Efficacy of Polyvalent Bacteriophage P-27/HP to Control Multidrug Resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> Associated with Human Infections

Emergence of multidrug resistant strains has created serious problem for safe eradication of Staphylococcus aureus infections. Therefore, there is an urgent need to develop novel antibacterial agents to control this pathogen. Bacteriophages kill bacteria irrespective of their antibiotic sensitivity and thus they can be used as potent prophylactic/therapeutic agent to treat such infections. Here, we report isolation of broad host range anti-staphylococcal lytic bacteriophage P-27/HP from sewage water. This phage was able to inhibit 17 of 28 (60%) human disease associated S. aureus isolates. In vitro studies revealed its strong lytic efficacy to diminish S. aureus 27/HP population (c.f.u.) by more than 5.0 logs (P < 0.0001) (equivalent to 99.99%) in 3 h at 0.01 MOI. In vivo lytic efficacy analysis showed that a single subcutaneous injection of phage P-27/HP (10⁷ p.f.u.) was sufficient to protect S. aureus 27/HP infected (5 × 10⁸ c.f.u.) mice from bacteremia and subsequent death. A considerable decline of more than 6 logs (99.9%) in splenic S. aureus 27/HP c.f.u. count was noted at the 3 days of phage treatment. In conclusion, our results suggest that phage P-27/HP is polyvalent in nature and has high-lytic potential towards S. aureus, thus, a therapy employing this phage would be efficacious to control S. aureus infections.     

Aerobic heterotrophic bacteria and petroleum-utilizing bacteria from cow dung and poultry manure

In this study, enumeration and identification of total aerobic heterotrophic bacteria and petroleum-utilizing bacteria as well as the degradative potential of petroleum-utilizing bacterial isolates were carried out. The average counts of total aerobic heterotrophic bacteria in cow dung and poultry manure were 74.25 × 10⁵ c.f.u. g⁻¹ and 138.75 × 10⁵ c.f.u. g⁻¹ respectively. Acinetobacter sp, Bacillus sp, Pseudomonas sp, and Serratia spp. occurred as aerobic heterotrophs in both cow dung and poultry manure. However, Alcaligenes spp. occurred only in cow dung while, Flavobacterium sp, Klebsiella sp, Micrococcus sp, and Nocardia spp. occurred only in poultry manure as aerobic heterotrophs. The average counts of petroleum-utilizing bacteria in cow dung and poultry manure were 9.25 × 10⁵ c.f.u. g⁻¹ and 17.25 × 10⁵ c.f.u. g⁻¹ respectively. Pseudomonas spp. occurred as petroleum utilizer in both cow dung and poultry manure. However, Bacillus spp. occurred only in cow dung while Acinetobacter spp. and Micrococcus spp. occurred only in poultry manure as petroleum utilizers. Relative abundance of petroleum utilizers in total aerobic heterotrophs ranged from 6.38% to 20.00% for cow dung and from 9.38% to 17.29% for poultry manure. Introduction of pure cultures of petroleum-utilizing bacteria from cow dung and poultry manure into sterile oil-polluted soil revealed oil degradation in one week period.     

Microbial Diversity in Ngari,Hentak and Tungtap,Fermented Fish Products of North-East India

Ngari, hentak and tungtap are traditional fermented fish products of North-East India. Eighteen samples of ngari, hentak and tungtap were collected and were analysed for microbial load. Lactic acid bacteria, endospore-forming rods, yeasts and aerobic mesophilic counts ranged from 4.0 to 7.2, 3.3–4.6, <1–3.5 and 4.3–7.3 log c.f.u./g, respectively. Lactic acid bacteria were identified as Lactococcus lactis subsp. cremoris, Lactococcus plantarum, Enterococcus faecium, Lactobacillus fructosus, Lactobacillus amylophilus, Lactobacillus coryniformis subsp. torquens and Lactobacillus plantarum. Endospore-forming rods were identified as Bacillus subtilis and Bacillus pumilus, aerobic coccal strains were identified as Micrococcus. Yeasts were identified as species of Candida and Saccharomycopsis. Pathogenic contaminants were detected in all samples, however, none of the sample contained more than 10² c.f.u./g of Bacillus cereus, 10³ c.f.u./g of Staphylococcus aureus and enterobacteriaceae population, respectively. Enzymatic and antimicrobial activities of the isolates were tested. None of the strains produced biogenic amines in the method applied. Most strains of LAB had a high degree of hydrophobicity, indicating their ‘probiotic’ characters. This study has demonstrated the microbial diversity within the species of lactic acid bacteria, Bacillus and yeasts. This revised version was published online in July 2006 with corrections to the Cover Date.     

An engineered Bacillus thuringiensis strain with insecticidal activity against Scarabaeidae (Anomala corpulenta) and Chrysomelidae (Leptinotarsa decemlineata and Colaphellus bowringi)

A genetically-engineered Bacillus thuringiensis (Bt) strain, 3A-HBF, with a broad insecticidal spectrum was constructed by introducing the recombinant plasmid pSTK-3A containing cry3Aa7 into the wild-type Bt strain HBF-1 containing the cry8Ca2 gene. The Cry3Aa7 protein produced by strain 3A-HBF was verified by SDS-PAGE and Western blotting. Flat rectangular crystals of Cry3Aa7 protein were observed besides spherical crystals (Cry8Ca2). The plasmid pSTK-3A was stable when strain 3A-HBF was grown in medium without antibiotics. The growth rate of 3A-HBF was not significantly different from that of the recipient strain, HBF-1. Strain 3A-HBF showed toxicity against two families of pests, Scarabaeidae and Chrysomelidae pests, which are susceptible to Cry8Ca (Anomala corpulenta) and Cry3Aa (Leptinotarsa decemlineata and Colaphellus bowringi). The 50% lethal concentrations of 3A-HBF against A. corpulenta, L. decemlineata and C. bowringi were 0.730 × 10⁸ c.f.u./g dry soil, 1.74 μg/ml and 1.15 μg/ml, respectively.     

Production,formulation and antagonistic activity of the biocontrol like-yeast <Emphasis Type="Italic">Aureobasidium pullulans</Emphasis> against <Emphasis Type="Italic">Penicillium expansum</Emphasis>

Aureobasidium pullulans (de Bary) Arnaud (Ach 1-1) was grown in a glucose fed-batch fermentor to 106 g dry wt l⁻¹ in 48 h. The cells were dried in a fluidized bed dryer with a final viability of 62%. After 7 months at 4°C, the viability was 28% of the initial value (= 2.3 × 10¹⁰ c.f.u. g⁻¹ dry matter). A protection level of 89% was achieved with the biomass preparation at 1 × 10⁸ c.f.u. ml⁻¹ after 28 and 7 days for apples stored respectively at 5 and 25°C against Penicillium expansum. Our process is suitable to produce large quantities of the strain Ach 1-1 as biological control agent for apple preservation.     

Identification of <Emphasis Type="Italic">Aeromonas salmonicida</Emphasis> in European perch from North Lithuanian rivers during mass mortalities in 2008

The investigation was carried out to ascertain the cause of mass mortalities in European perch (Perca fluviatilis L.) in North Lithuanian rivers in 2008 that reached a maximum in mid-autumn at a water temperature of 6°C. Marked changes were detected in morphophysiological parameters (spleen-, gill-, liver- and heart-somatic indices), which corresponded to changes in haematological parameters (leukocyte count was significantly elevated or reduced) of the infected fish. The viable bacterial counts in the gill, liver and kidney of infected live fish samples ranged from 2.3±0.3 × 10³ to 6.3×00B1;0.4 × 10³ c.f.u./g. The BOX-PCR fingerprinting technique was used for characterization and identification of the isolated bacterial strains from liver and kidney of infected perch. The bacterial isolates were identified by sequence analysis of the 16S rRNA gene. Pathogenic Aeromonas salmonicida subsp. salmonicida has been identified as a possible causative agent of the observed mortality in European perch.     

Microbial flora and some chemical properties of ersho,a starter for teff (Eragrostis tef) fermentation

Ersho is a clear, yellow liquid that accumulates on the surface of fermenting teff-flour batter and is collected to serve as an inoculum for the next fermentation. The pH of ersho samples was about 3.5 and titratable acidity ranged between 3.1% and 5.7%. The mean aerobic mesophilic counts from four households varied between 6.9×10⁶ and 1.3×10⁸ c.f.u./ml and the aerobic bacterial flora consisted of Bacillus spp. Mean yeast counts ranged between 5.2×10⁵ and 1.8×10⁶ c.f.u./ml and comprised, in order of abundance, Candida milleri, Rhodotorula mucilaginosa, Kluyveromyces marxianus, Pichia naganishii and Debaromyces hansenii. Candida milleri was the most dominant isolate in all samples. About 90% of the teff flour samples had aerobic mesophilic counts 10⁵ c.f.u./g and Gram-positive bacteria constituted about 71% of the total isolates. About 80% of samples had Enterobacteriaceae counts of 10⁴ c.f.u./g.M. Ashenafi is with the Department of Basic Sciences, Awassa College of Agriculture, Addis Ababa University, PO Box 5, Awassa, Sidamo, Ethiopia.     

Microbial load,incidence and antibiotic resistance of some disease-causing microorganisms on raw food items in consumed Ethiopia

Summary A total of 708 strains belonging to different bacterial genera were isolated from fresh tomatoes, eggs, green peppers and fish sold in open markets in Awassa, Ethiopia. They mainly consisted of coliforms (54%),Staphylococcus aureus (26%) andProteus (16%).Shigella was isolated from 26% of the tomato specimens. Total counts for the various food items varied between 2.4 × 10⁶ c.f.u./g and 9.5 × 10⁶ c.f.u./g. Most probable number of total coliform and faecal coliform ranged from 1.5 × 10²/g to 3 × 10²/g and 37/g to 70/g. Less than 50% of the isolated 522 Gram-negative rods were susceptible to ampicillin and cephalothin and only 14 isolates were susceptible to all drugs tested.Proteus showed higher rates of resistance to ampicillin, carbenicillin and cephalothin (75%). Resistance to these drugs was also relatively higher inEscherichia coli (50%). Sixty-eight of the 186S. aureus isolates were sensitive to all the drugs tested and susceptibility to tetracycline and chloramphenicol was relatively lower (70%).

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Charge microbienne, incidence et résistance aux antibiotiques de quelques micro-organismes dans des aliments destinés à être consommés crus en Ethiopie

Résumé On a isolé un total de 708 souches appartenant à divers genres bactériens de tomates fraîches, d'oeufs, de poivrons verts et de poissons vendus sur les marchés ouverts en Awassa, Ethiopie. Parmi celles-ci, on a trouvé 26% deStaphylococcus aureus et 16% deProteus. On a isolé desShigella dans 26% des tomates testées. L'énumération totale des germes dans divers aliments a donné des valeurs situées entre 2.4 × 10⁶ c.f.u./g et 9.5 × 10⁶ c.f.u./g. Le nombre le plus probable de coliformes totaux et de coliformes fécaux variait entre 1.5 × 10²/g et 3 × 10²/g et entre 37/g et 70/g. Moins de 50% des 522 souches isolées à forme de bâtonnets étaient sensibles à l'ampicilline et à la céphalothine et seules 14 souches isolées étaient sensibles à toutes les drogues testées. LesProteus ont montré davantage de résistance à l'ampicilline, la carbénicilline et la céphalothine (75%). La résistance à ces drogues était aussi relativement plus élevée chezEscherichia coli. Soixante huit de 186 souches isolées deS. aureus étaient sensibles à toutes les drogues testées et leur sensibilité à la tétracycline et au chloramphenicol était relativement plus faible (70%).

     

Effects of Acetic Acid on the Regrowth of Heterotrophic Bacteria in the Drinking Water Distribution System

Summary Three laboratory-scale water pipe systems were set up to study the effects of adding two levels of acetic acid (10 and 50 μg acetate eq-C l⁻¹) on the bacterial regrowth in water pipes. The results of the water pipe test showed that nearly all carbon in the acetic acid could be readily utilized by bacteria and resulted in an increase in biomass concentration. The maximum heterotrophic plate counts in biofilm were equal to 3.5 × 10⁴, 8.9 × 10⁵ and 2.9 × 10⁷ c.f.u. cm⁻² while the maximum heterotrophic plate counts of free bacteria were equal to 1.2 × 10³, 5.0 × 10³ and 6.8 × 10⁴ c.f.u. ml⁻¹ for the blank and with addition of 10 and 50 μg acetate eq-C l⁻¹. These results showed that addition of acetic acid to drinking water has a positive effect on the assimilable organic carbon content of drinking water and bacterial regrowth in the distribution system. This effect is enhanced with addition of high-level acetic acid. Batch tests were also conducted using water samples collected from a Taiwanese drinking water distribution system. The bacterial regrowth potentials of the blank were equal to 4.3 × 10³, 1.5 × 10⁴, 4.9 × 10⁴ and 7.5 × 10⁴ c.f.u. ml⁻¹ for water samples collected from treatment plant effluent, commercial area, mixed area, and residential area, respectively. These results showed that the biological stability of drinking water is the highest in treatment plant effluent, followed by distributed water of the commercial area, distributed water of the mixed area, and then the distributed water of residential area.