Aerobic Spore-forming Bacteria Research Articles

Characterization of toxigenic genes of Bacillus cereus strains isolated from different spices sold in Algeria.

Many cooked foods are prepared with spices and dried herbs; these can be contaminated by several types of microorganisms, including aerobic spore-forming bacteria. The Bacillus cereus group is very widespread in nature and is known among the common food contaminants. They are involved in food poisoning, causing two types of syndromes, diarrheal and emetic. The aims of the present work were to determine the prevalence of toxigenic Bacillus cereus spores in spices and herbs marketed in the Laghouat area and to identify their toxigenic genes via PCR. Among the 191 samples, 14.13% were determined to be B. cereus, with concentrations ranging from 2.52 to 5.82 log cfu/g, where the highest level of contamination was observed in allspice and ginger. Moreover, entFM (100%), nhe (88.23%) and cytK (70.58%) were the most frequently identified toxin genes, whereas hbl (23.52%) was less common, and no emetic toxin-encoding gene (cesB) was found in any of the samples. Considering the results of the present study, the B. cereus microbial load and toxin gene profiles of spices show that spices have potential for public health in Algeria. In this context, it is crucial to guarantee the microbiological safety of spices by respecting good hygiene practices, eliminating bacterial spores and toxin production via sterilization and using appropriate packaging for these products.

Potential effect of monolaurin and nisin on the growth of three aerobic spore-forming bacteria isolated from Tunisian milk

Antibacterial capacities of monolaurin and nisin on the growth of vegetative cells of three sporeforming bacteria isolated from cow milk collected from Tunisia (Terribacillus aidingensis, Bacillus sporothermodurans and Paenibacillus sp and) were evaluated. Nisin or monolaurin, at different concentrations, can inhibit the growth of three Bacillus. 500 IU/mL and 25 μg/mL corresponded to the minimum inhibitory concentrations of nisin and monolaurin, respectively. The growth of the tested species in the presence of sub-lethal concentrations of monolaurin and nisin was monitored in UHT milk for 7 days at three different temperatures (4°C, 37°C and 25°C). Nisin (250 IU/mL) was able to induce an immediate reduction (after 3 h of incubation at 4°C) of Bacillus species growth (5 log for B. sporothermodurans, 4 log for Paenibacillus sp and 3.2 log for T. aidingensis). Also, monolaurin (25 µg/mL) reduce the growth, at 4°C of B. sporothermodurans by 5 log, Paenibacillus sp by 3 log and T. aidingensis by 4 log, after 3 h of incubation. In the presence of nisin or monolaurin, it can be observed a low reduction in cell growth followed by a rapid regrowth which can attenuate 10 log, at 37°C and 25°C. The obtained data elucidate the potency of nisin and monolaurin, and the application of these antimicrobial agents at low temperatures (4°C) to decontaminate any product from these spoilage bacteria.

Comparative Analysis of Alkaline Phosphatase Activity and Aerobic Spore-Forming Bacteria in Pasteurized Milk

Comparative Analysis of Alkaline Phosphatase Activity and Aerobic Spore-Forming Bacteria in Pasteurized Milk

Review on Microbiology of Cereal-Based Spontaneously Fermented Foods and Beverages

Spontaneously fermented cereal-based foods and beverages are fermented by diverse arrays of microorganisms which play significant roles at different stages of fermentation. The aim of this review is to summarize the scientific data on the microbiology of cereal-based spontaneously fermented foods and beverages. Yeasts are a large group of beneficial bacteria in food fermentation followed by lactic acid bacteria. Molds also play an important role in the production of various foods and non-food products. The possible functions of yeast in the fermentation of carbohydrates are aroma production, stimulation of lactic acid bacteria and degradation of mycotoxin. However, this review results reveal that all yeasts and molds are not beneficial microbes. According to the reports of many researchers, <i>Enterobacteriaceae</i> and total coliforms are not persisted to the end of fermentation. However, aerobic mesophilic bacteria, staphylococcus spp., and aerobic spore-forming bacteria are persisted to the end of some fermented food and beverage products. In spontaneous fermentation, aerobic spore-forming bacteria play crucial role in the begging stage of fermentation to breakdown of complex structure of food composition which facilitates the environment for the yeast and lactic acid bacteria. This review concluded that spontaneous fermentation process reduces both pathogenic and spoilage microorganisms to non-significant level thus make the end product safe for consumption.

Aerobic spore-forming bacteria associated with ropy bread: Identification, characterization and spoilage potential assessment

Aerobic spore-forming (ASF) bacteria have been reported to cause ropiness in bread. Sticky and stringy degradation, discoloration, and an odor reminiscent of rotting fruit are typical characteristics of ropy bread spoilage. In addition to economic losses, ropy bread spoilage may lead to health risks, as virulent strains of ASF bacteria are not uncommon. However, the lack of systematic approaches to quantify physicochemical spoilage characteristics makes it extremely difficult to assess rope formation in bread. To address this problem, the aim of this study was to identify, characterize and objectively assess the spoilage potential of ASF bacteria associated with ropy bread.Hence, a set of 82 ASF bacteria, including isolates from raw materials and bakery environments as well as strains from international culture collections, were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and their species identity confirmed by 16S rRNA and gyrA or panC gene sequencing. A standardized approach supported by objective colorimetric measurements was developed to assess the rope-inducing potential (RIP) of a strain by inoculating autoclaved bread slices with bacterial spores. In addition, the presence of potential virulence factors such as swarming motility or hemolysis was investigated.This study adds B. velezensis, B. inaquosorum and B. spizizenii to the species potentially implicated of causing ropy bread spoilage. Most importantly, this study introduces a standardized classification protocol for assessing the RIP of a bacterial strain. Colorimetric measurements are used to objectively quantify the degree of breadcrumb discoloration. Furthermore, our results indicate that strains capable of inducing rope spoilage in bread often exhibit swarming motility and virulence factors such as hemolysis, raising important food quality considerations.

Особливості онтогенезу бацил впродовж розвитку від спори до вегетативної клітини

In the review, on the example of aerobic spore-forming bacteria, the problems of the development and ontogenesis of the bacterial cell are considered along with the possibilities of influencing the course of these processes. The characteristics of the main concepts "growth", "differentiation", and "development" as independent processes with their dynamic interrelation are presented. Attention is focused on the analysis of literature data on the peculiarities of vegetative cell development, starting from a spore in a dormant state and finishing with vegetative form formation. In particular, the mechanisms that maintain the spore dormant state and subsequent processes of activation, initiation, outgrowth, and vegetative cell formation are described. There are emphasized certain problems with research on the ontogenesis of bacterial cells due to the deficiency of appropriate methods, as well as the lack of a single opinion regarding individual stages of the development and vegetative form formation. It was concluded that the study of individual stages of the development of prokaryotes, which differ in spore-forming and non-spore-forming microorganisms, is still relevant. Knowledge of these processes will help scientists to develop mechanisms of influence on the ontogenesis of microorganisms.

Isolation of aerobic bacteria from abscesses and wounds in rabbits and antibiotic susceptibility testing of Staphylococcus spp. and Pseudomonas spp. isolates

BackgroundWound infections and abscesses in rabbits are a common problem in veterinary practice and their treatment can be challenging. Bacterial culture and antimicrobial susceptibility testing are important tools in these cases. While some studies are available on bacteria associated with wound infections and abscesses in rabbits, the numbers of animals included is generally limited. Monitoring of antibiotic resistance in clinical practice is also an important part of good antibiotic stewardship. Multiple antibiotic resistances are frequently encountered in Staphylococcus and Pseudomonas spp., two groups of bacteria commonly associated with abscesses in rabbits. MethodsThis retrospective study evaluated the results of aerobic bacterial cultures and antibiotic susceptibility tests in swabs from abscesses (A) (n=541) and from skin and wounds (SW) (n=308) submitted to a commercial diagnostic laboratory. Samples submitted between January 2017 and October 2020 were included in the study. Antimicrobial susceptibility testing for Staphylococcus and Pseudomonas spp. was evaluated and compared. ResultsIn A, 465 bacterial isolates including 35 different species were isolated while 394 bacterial isolates, including 35 different species, were found in SW. The most frequently isolated bacteria were Staphylococcus spp. (A = 21.7%, SW = 36.5%) followed by Pasteurella spp. in A (13.3%) and aerobic spore-forming bacteria in SW (11.2%). Pseudomonas spp. was the third most commonly isolated genus in both sample types (A = 11.6%, SW = 8.9%). Pseudomonas aeruginosa isolated from A were more often resistant to the tested antibiotics than other Pseudomonas species. These results support the need for bacterial culture and antibiotic susceptibility testing in small mammal practice.

Melissopalynological analysis and microbiological safety of fresh and market honey (Apis mellifera L. and Meliponulabeccarii L.) from Western Oromia, Ethiopia

Honey is a natural product that is made by bees from the nectar of flowering plants. There is a flora preference by bees. Like other foods ready to eat,honey can be prone to microbial contamination. Honey plant sources can be analyzed from the composition of pollen grains in honey samples. The objective of this study was to assess microbial safety and floral origin of the honey samples. For this study, honey samples were purchased from local market, and collected from hives (fresh honey) in Western Oromia. Floral analysis was determined using harmonized method of melissopalynology. Microbiological safety was assessed through the pour plate procedures from the first serial dilution on a total of 45 honey sample sizes.The melissopalynological analysis demonstrated that A. melliferahoney purchased from the market(AMMH) was considered a multi-floral type while A. mellifera fresh honey (AMFH) cropped directly from the hive and M.beccarii honey purchased from the market (MBMH) was dominated pollen from Coffee arabica (68 % of its pollen grain counted) and Guizotiascabra (50.53 % of its pollen grain counted) plant, respectively. The Aerobic mesophilic bacteria, Staphylococci, Yeast, Mould, and Aerobic spore-forming bacteria were found below the standard countable level (<30 cfu/plate) from A. mellifera and M.beccarii honey bought from the market, while A. mellifera honey collected directly from the hive became free of any microbial contamination. C.arabica and G.scabra are major honey plants and their honey can be harvested in February and October, respectively. Furthermore, Vernoniaamygdalina, Eucalyptus spp, Combretummolle, Trifoliumruppelianum, and Syzgiumguineense were honey plants analyzed from multifloral market honey even though, their pollen dominance varies. M. beccarii visits herbaceous flora whilst A. mellifera visits all floral types. The level of contamination of the honey samples from the study area was very low showing its safety.

Phenotypic, genotypic, and resistome of mesophilic spore-forming bacteria isolated from pasteurized liquid whole egg

The production of whole-liquid eggs is of significant economic and nutritional importance. This study aimed to assess the phenotypic and genotypic diversity of mesophilic aerobic spore-forming bacteria (n = 200) isolated from pasteurized whole liquid egg and liquid egg yolk. The majority of the isolates were identified as belonging to the genera Bacillus (86 %), followed by Brevibacillus (10 %) and Lysinibacillus (4 %). For the phenotypic characterization, isolates were subjected to various heat shocks, with the most significant reductions observed at 80 °C/30 min and 90 °C/10 min for isolates recovered from raw materials. On the other hand, the decrease was similar for isolates recovered from raw material and final product at 100 °C/5 min and 110 °C/5 min. Genotypic genes related to heat resistance (cdnL, spoVAD, dacB, clpC, dnaK, and yitF/Tn1546) were examined for genotypic characterization. The dnaK gene showed a positive correlation with the highest thermal condition tested (110 °C/5 min), while 100 °C/5 min had the highest number of positively correlated genes (clpC, cdnL, yitF/Tn1546, and spoVAD). Whole Genome Sequencing of four strains revealed genes related to sporulation, structure formation, initiation and regulation, stress response, and DNA repair in vegetative cells. The findings of this study indicate that these mesophilic aerobic spore-forming bacteria may adopt several strategies to persist through the process and reach the final product. As the inactivation of these microorganisms during egg processing is challenging, preventing raw materials contamination and their establishment in processing premises must be reinforced.

Farmed Insect Frass as a Future Organic Fertilizer

The aim of this incubation experiment was to evaluate the effect of Tenebrio molitor L. frass on selected chemical and microbiological properties of acid peat. The optimal rate of mealworm frass in the substrate for growing ornamental trees and shrubs was determined. Acid peat was fertilized with frass or urea at five nitrogen (N) rates: 0, 50, 100, 200, and 400 mg dm−3. Mineral N content and electrical conductivity increased, and calcium content decreased in peat with increasing N rates. Similarly to urea, frass increased the ammonification rate at the beginning of incubation and the nitrification rate from the second week of incubation. Higher frass rates increased the abundance of plant-available nutrients (N, P, Mg, K, and Na) in acid peat. Frass undesirably decreased the counts of bacteria with antagonistic activity against soil-borne plant pathogens. Regarding the abundance of functional genes, the optimal N rate was 100 mg dm−3, which promoted the growth of N-fixing and chitinolytic bacteria. Higher N rates promoted the development of aerobic spore-forming bacteria, which produce antibiotics that can be used as biocontrol agents. Moderate fertilizer rates contributed to N accumulation in bacterial biomass. These preliminary findings, which indicate that insect frass can partially replace mineral fertilizers, are promising and can be used in pot and field experiments testing various plant species.

Modern research on the dynamics of soil microbial communities in soils of Azerbaijan

In soils with minimal anthropogenic load the following were found B.cereus, B.muralis, B. Cibi, B. Simplex, Br.laterosporus, and in soils with high anthropogenic load the following were found B. cibi, B.cereus, P.lautus, B. pseudomycoides, B. niacini, B. mojavensis, B. agaradhaerens, B.arsenicus, B.firmus, B.licheniformis, B.atrophaeus, P. polymyxa, B.acidicola, B.drentensis, Br.laterosporus, B.subtilis. In anthropogenically transformed soils, the number of common bacilli species was reduced, while at each point species were found that were not found at other points. The proportion of microorganisms that utilize natural substrates is sharply reduced - in urban soils the number of aerobic spore-forming bacteria of the order Bacillales decreases several times.

Production and Biochemical, Phytochemical, Sensory, and Microbiological Characterization of “Bili-Bili” and “Cochette,” Two Traditional Beers Produced from Sorghum (Sorghum bicolor) and Rice (Oryza sativa)

“Bili-bili” and “cochette” are two traditional beers made from sorghum and rice, respectively. Despite the socioeconomical interest of these drinks, limited studies have been conducted to elucidate their quality and safety. To fill these gaps, the quality features and the safety status of both beers were assessed. After a reasoned field survey, the samples from various locations in Maroua were collected and analysed using referenced methods. The field survey revealed that both beers were produced under unsanitary conditions using rudimentary procedures. Mean values of pH (2.99 and 3.58), TTA (1.1 and 0.9%), alcohol (3.8 and 2.5%), DM (6.4 and 11.1%), TSS (6.94 and 6.18 °Brix), proteins (0.54 and 0.71 g/100 mL), amino acids (0.30 and 0.38 g/100 mL), and ash (1.52 and 0.51%) were recorded in “bili-bili” and “cochette,” respectively. Similarly, the TPC, TFC, and carotenoid content of 325.5 and 352.4 mgGAE/100 mL, 314.4 and 278.9 mgQE/100 mL, and 95.4 and 89.4 mg/100 mL were noted in both beers, respectively. “Bili-bili” and “cochette” exhibited free radical scavenging activity of 42.4 and 36.7% and reducing power of 87.3 and 119.5 mgTE/100 mL, respectively. Overall acceptability ratings ranged from 5.6 to 7.5 and from 6.7 to 6.9 for “bili-bili” and “cochette,” respectively. No pathogen was detected, but the presence of total aerobic bacteria, fungi, coliforms, and aerobic spore-forming bacteria flora above the recommended limits made both beers unsafe and potentially harmful for consumers. Given all above, improving of the production scheme and microbiological quality of the two traditional beers are required to ensure the safety of consumers.

Identification and Characterisation of Spore-Forming Bacteria in Bovine Raw Milk Collected from Four Dairy Farms in New Zealand

Contamination of milk and dairy products with pathogenic and spoilage bacteria may result in huge economic loss due to recalls of products. This study aimed to identify spore-forming bacteria from raw milk and characterise those for toxin production and their spoilage ability. Bovine raw milk collected from dairy farms in the Manawatu region of the North Island of New Zealand was tested for the presence of both aerobic and anaerobic spore-forming bacteria using standard culture-based techniques, as well as genomic analysis. The spore-forming bacteria were investigated for the presence of toxin genes and their spoilage potential. A low number of aerobic spore-forming bacteria were detected in raw-milk samples collected from the four farms in summer and winter. The 16S rRNA sequence types similar to important food spoilage bacteria like C. beijerinckii, C. sporogenes, B. licheniformis and members of the Paenibacillus genus, as well as potentially toxigenic bacteria such as B. cereus and C. perfringens were isolated. Genes responsible for important toxin production were present in some of the tested spore-forming bacteria. This pilot study highlights the presence of various spoilage and pathogenic spore-forming bacteria in raw milk from these farms. A low number of spore-forming bacteria indicates the implementation of good hygienic farm practices and management to reduce the contamination of raw milk with spore-forming bacteria.

Upflow Direct Filtration and Downflow Direct Filtration: A Comparison with Emphasis on the Removal of Aerobic Spore-Forming Bacteria and Fluorescent Microspheres as Surrogates for Cryptosporidium Oocyst Removal

Monitoring the removal of Cryptosporidium oocysts in water treatment plants is a challenge, particularly in developing countries, and the use of surrogates has proven necessary. Two surrogates already investigated and recognized by the scientific community are aerobic spore-forming bacteria (ASFB) and fluorescent polystyrene microspheres (ME). In this context, this study evaluates and compares the performance of upflow direct filtration and downflow direct filtration, with emphasis on the removal of ASFB and ME as a surrogate of Cryptosporidium oocyst removal. Pilot filtration experiments were carried out with low turbidity Paranoá Lake water (average—3.7 NTU) with ASFB varying from 7 × 102 to 5.5 × 103 CFU/100 mL. In some experiments, water was spiked with microspheres, aiming for 105 ME/L. Two operational conditions were evaluated: (1) filters operating with the same filtration rate (5 m/h); (2) filters operating with the same flow rate of 59.41 L/h. In general, the downflow filter presented a slightly higher removal efficiency than the upflow filter but the differences were not considered statistically significant in most cases. The removal efficiency of microspheres (3 to 5 log) was higher than that of ASFB (1.2 to 4 log) in both filters, corroborating that the removal of ASFB is a conservative surrogate of the removal of Cryptosporidium.

Microbacterium represents an emerging microorganism of concern in microfiltered extended shelf-life milk products

Growing interest in the manufacture of extended shelf-life (ESL) milk, which is typically achieved by a high temperature treatment called ultra-pasteurization (UP), is driven by distribution challenges, efforts to reduce food waste, and more. Even though high temperature, short time (HTST) pasteurized milk has a substantially shorter shelf-life than UP milk, HTST milk is preferred in the United States because consumers tend to perceive UP milk as less desirable due to the "cooked" flavor associated with high-temperature processing. While ESL beyond 21 d may be possible for HTST, the survival and outgrowth of psychrotolerant aerobic spore-forming bacteria can still be a limitation to extending shelf-life of HTST milk. Microfiltration (MF) is effective for reducing vegetative microorganisms and spores in raw milk, but it is unclear what the effects of membrane pore size, storage temperature, and milk type (i.e., skim vs whole) are on the microbial shelf-life of milk processed by both MF and HTST pasteurization. To investigate these factors, raw skim milk was MF using different pore sizes (0.8 μm or 1.2 μm), and then MF skim milk and standardized whole milk [MF skim with heat-treated (85°C for 20 s) cream] were HTST pasteurized at 75°C for 20 s. Subsequently, milk was stored at 3, 6.5, or 10°C and total bacteria counts were measured for up to 63 d. An ANOVA indicated that mean bacterial concentrations between storage temperatures were significantly different from each other, with mean maximum observed concentrations of 3.67, 5.33, and 8.08 log10 cfu/mL for storage temperatures 3, 6.5, and 10°C, respectively. Additionally, a smaller difference in mean maximum bacterial concentrations throughout shelf-life was identified between pore sizes (<1 log cfu/mL), but no significant difference was attributed to milk type. An unexpected outcome of this study was the identification of Microbacterium as a major contributor to the bacterial population in MF ESL milk. Microbacterium is a psychrotolerant, thermoduric gram-positive, non-spore forming rod with a small cell size (∼0.9 μm length and ∼0.3 μm width), which was able to permeate the membranes used in this study, survive HTST pasteurization, and then grow at refrigeration temperatures. While spores continue to be a key concern for the manufacture of MF, ESL milk, our study demonstrates the importance of other psychrotolerant, thermoduric bacteria such as Microbacterium to these products.

Influence of high-pressure homogenization treatments combined with lysozyme activated packaging on microbiological and technological quality of vegetable smoothie during shelf life

In this study the effect of a high pressure homogenization (HPH) treatment, alone and in combination with a biodegradable polylactic acid (PLA) packaging activated with lysozyme on a smoothie made with apple, carrot and rice beverage was evaluated. In particular, the effects of treatments on the microbiological and quality characteristics of smoothies were assessed during storage at 4 and 10 °C in climatic chambers at RH of 50 %. Obtained results showed the efficacy of HPH at 100 MPa to reduce the initial cell load of mesophilic and psycothrophic aerobic bacteria, lactic acid bacteria, yeasts and aerobic spore-forming bacteria. Moreover, the combination of HPH and lysozyme activated packaging resulted in an increased antimicrobial effect against mesophilic, psychrotrophic and lactic acid bacteria during product storage at 10 °C. Yeast and spore forming bacteria showed, instead, lower cell loads independently from samples. Samples packed in active packaging, both treated and not, showed a better color retention in terms of lightness and red index, compared to control (C) ones both when stored at 4 and 10 °C. The HPH treated smoothies, both activated and not, at 4 °C remained microbiologically stable for more than 20 days. However, the use of active packaging has also allowed to reduce the microbiological proliferation during storage at 4 °C. Moreover, few differences in color indices were highlighted between samples stored at 4 °C. Obtained results indicate that the combination of HPH treatment and active packaging may represent a useful strategy to increase safety and shelf-life of vegetable smoothies.

Бактерии р. Bacillus – антагонисты патогенов аквакультуры

Due to the increasing resistance of aquaculture pathogens to antibiotics, the search for new approaches to the control of infectious diseases is of particular importance. A promising approach is the development of probiotic preparations based on antagonist bacteria. The paper studied the antagonistic activity of Bacillus strains that are promising for developing probiotics for fish. The strains were isolated from the bottom sediments of the Don River in the area of the Donskoy Fish Reserve, as well as from the intestines of cyprinids of natural populations, namely, from silver carp (Cyprinus gibelio), roach (Rutilus heckelii), and bream (Abramis brama). The inhibitory effect of Bacillus strains against causative agents of aeromonosis, pseudomonosis and bacterial hemorrhagic septicemia was studied by the method of delayed antagonism. Aeromonas and Pseudomonas species pathogenic for fish were isolated from sturgeons, cyprinids and salmons grown in pond farms and recirculating aquaculture systems in the Krasnodar Region, Rostov Region and the Republic of Adygea. It has been shown that sensitivity to the action of bacilli is not only a species, but also a strain-specific trait among bacterial pathogens of fish. Five strains were identified that exhibited the maximum antagonistic activity against the studied pathogens, and their species identification was carried out by molecular biological and mass spectrometric methods. It has been established that the most pronounced antagonism is exhibited by strains of bacilli isolated from bottom sediments. The growth of all studied strains of fish pathogenic species of Pseudomonas was suppressed only the B. subtilis R4 strain. Thus, strains of aerobic spore-forming bacteria suitable for the creation of probiotic preparations for aquaculture objects have been isolated and characterized.

Shelf-life storage temperature has a considerably larger effect than high-temperature, short-time pasteurization temperature on the growth of spore-forming bacteria in fluid milk

In the absence of postpasteurization contamination, psychrotolerant, aerobic spore-forming bacteria that survive high-temperature, short-time (HTST) pasteurization, limit the ability to achieve HTST extended shelf-life milk. Therefore, the goal of the current study was to evaluate bacterial outgrowth in milk pasteurized at different temperatures (75, 85, or 90°C, each for 20 s) and subsequently stored at 3, 6.5, or 10°C. An initial ANOVA of bacterial concentrations over 14 d of storage revealed a highly significant effect of storage temperatures, but no significant effect of HTST. At d 14, average bacterial counts for milk stored at 3, 6.5, and 10°C were 1.82, 3.55, and 6.86 log10 cfu/mL, respectively. Time to reach 1,000,000 cfu/mL (a bacterial concentration where consumers begin to notice microbially induced sensory defects in fluid milk) was estimated to be 68, 27, and 10 d for milk stored at 3, 6.5, and 10°C, respectively. Out of 95 isolates characterized with rpoB allelic typing, 6 unique genera, 15 unique species, and 44 unique rpoB allelic types were represented. The most common genera identified were Paenibacillus, Bacillus, and Lysinibacillus. Nonmetric multidimensional scaling identified that Bacillus was significantly associated with 3 and 10°C, whereas Paenibacillus was consistently found across all storage temperatures. Overall, our data show that storage temperature has a substantially larger effect on fluid milk shelf life than HTST and suggests that abuse temperatures (e.g., storage at 10°C) allow for growth of Bacillus species (including Bacillus cereus genomospecies) that do not grow at lower temperatures. This indicates that stringent control of storage and distribution temperatures is critical for producing extended shelf-life HTST milk, particularly concerning new distribution pathways for HTST pasteurized milk (e.g., electronic commerce), and when enhanced control of spores in raw milk is not feasible.

Microbiological status of natural and anthropogenic soils of the Taganrog Bay coast at different levels of combined pollution with heavy metals and PAHs.

The effect of heavy metals (HMs) and polycyclic aromatic hydrocarbons (PAHs) pollution on the microbiological status of soils on the coast of the Taganrog Bay and adjacent areas was studied. The content of total and exchangeable forms of HMs, the content of 16 priority PAHs and the abundance of several groups of culturable microorganisms was determined, namely copiotrophic, prototrophic, aerobic spore-forming bacteria, actinomycetes, molds and yeasts. The content of total and exchangeable forms of HMs in urban coastal soils in industrial zone significantly exceeded that in non-urban soils. The maximum concentrations of total forms of Mn, Cr, Ni, Cu, Zn, Pb and Cd are 1821, 871, 143, 89, 1390, 317 and 10mg/kg, respectively. The median value of the total content of 16 PAHs in urban soils is 3 times higher than in the soils of natural areas and reached 4309ng/g. The lowest numbers of copiotrophic bacteria, prototrophic bacteria and aerobic spore-forming bacteria were found in the soils of industrial zone: 6.8, 13.8 and 0.63million CFUg-1 dry soil, respectively. The largest numbers of copiotrophic bacteria, prototrophic bacteria and aerobic spore-forming bacteria were recorded in the soils of natural areas-72.5, 136 and 5.73million CFUg-1 dry soil, respectively. It was found that the abundance of copiotrophs, prototrophs, and aerobic spore-forming bacteria is more affected by the urbanization of coastal soils including the pollution of HMs and PAHs. Other groups of microorganisms (actinomycetes, molds and yeasts) turned out to be more resistant to anthropogenic factors.

Alkaline protease production by thermotolerant Bacillus sp. KU-K2, from non-rubber skim latex through the non-sterile system and its enzymatic characterization

The non-rubber skim latex was used as a substrate for alkaline protease production under a non-sterile system. Among 108 isolates of aerobic spore-forming bacteria, the isolate of KU-K2 was selected as a potent bacterial strain for alkaline protease production. The 16s rRNA gene sequence analysis showed that the KU-K2 strain was closely related to the B. licheniformis with 99.39% similarity. The alkaline protease production from non-rubber skim latex serum was optimized using the Plackett-Burman design (PB) and response surface methodology based on central composite design (CCD). The results showed that the optimized medium consisted of 9.82 g/L skim milk, 0.97 g/L glucose, 0.82 g/L K2HPO4, and 0.02 g/L CoCl2·6H2O gave the highest enzyme production at 6.14 ± 0.13 U/mL. The upscale production in the 5.0 L stirrer fermenter was achieved at 6.78 ± 0.32 U/mL when incubated at 37 °C, 250 rpm with 2.0 vvm of aeration rate for 48 h, which increased by 2.90 folds compared to the un-optimized medium. The optimal pH and temperature for alkaline protease activity were 10.0 and 60 °C, respectively. Zn2+, Fe3+, and Hg2+ metal ions strongly inhibited the crude alkaline protease. This study showed the potential of thermotolerant Bacillus sp. For alkaline protease production using non-rubber skim latex as a substrate which could be applied to reduce the industrial waste from latex industries.