Kefir Research Articles

Integration of Meta-Multi-Omics Data Using Probabilistic Graphs and External Knowledge.

Multi-omics has the promise to provide a detailed molecular picture of biological systems. Although obtaining multi-omics data is relatively easy, methods that analyze such data have been lagging. In this paper, we present an algorithm that uses probabilistic graph representations and external knowledge to perform optimal structure learning and deduce a multifarious interaction network for multi-omics data from a bacterial community. Kefir grain, a microbial community that ferments milk and creates kefir, represents a self-renewing, stable, natural microbial community. Kefir has been shown to have a wide range of health benefits. We obtained a controlled bacterial community using the two most abundant and well-studied species in kefir grains: Lentilactobacillus kefiri and Lactobacillus kefiranofaciens. We applied growth temperatures of 30 °C and 37 °C and obtained transcriptomic, metabolomic, and proteomic data for the same 20 samples (10 samples per temperature). We obtained a multi-omics interaction network, which generated insights that would not have been possible with single-omics analysis. We identified interactions among transcripts, proteins, and metabolites, suggesting active toxin/antitoxin systems. We also observed multifarious interactions that involved the shikimate pathway. These observations helped explain bacterial adaptation to different stress conditions, co-aggregation, and increased activation of L. kefiranofaciens at 37 °C.

Enhanced probiotic potential of Lactobacillus kefiranofaciens OSU-BDGOA1 through co-culture with Kluyveromyces marxianus bdgo-ym6.

Due to the increasing consumer demand for the development and improvement of functional foods containing probiotics, new probiotic candidates need to be explored as well as novel means to enhance their beneficial effects. Lactobacillus kefiranofaciens OSU-BDGOA1 is a strain isolated from kefir grains that has demonstrated probiotic traits. This species is the main inhabitant of kefir grains and is responsible for the production of an exopolysaccharide (EPS) whit vast technological applications and potential bioactivities. Research has shown that interkingdom interactions of yeast and lactic acid bacteria can enhance metabolic activities and promote resistance to environmental stressors. Comparative genomic analyses were performed to distinguish OSU-BDGOA1 from other strains of the same species, and the genome was mined to provide molecular evidence for relevant probiotic properties. We further assessed the cumulative effect on the probiotic properties of OSU-BDGOA1 and Kluyveromyces marxianus bdgo-ym6 yeast co-culture compared to monocultures. Survival during simulated digestion assessed by the INFOGEST digestion model showed higher survival of OSU-BDGOA1 and bdgo-ym6 in co-culture. The adhesion to intestinal cells assessed with the Caco-2 intestinal cell model revealed enhanced adhesion of OSU-BDGOA1 in co-culture. The observed increase in survival during digestion could be associated with the increased production of EPS during the late exponential and early stationary phases of co-culture that, by enhancing co-aggregation between the yeast and the bacterium, protects the microorganisms from severe gastrointestinal conditions as observed by SEM images. Immune modulation and barrier function for recovery and prevention of flagellin-mediated inflammation by Salmonella Typhimurium heat-killed cells (HKSC) in Caco-2 cells were also measured. OSU-BDGOA1 in mono- and co-culture regulated inflammation through downregulation of pro-inflammatory cytokine expression and increased membrane barrier integrity assessed by TEER, FD4 permeability, and expression of tight junctions. The results of the study warrant further research into the application of co-cultures of yeast and LAB in functional probiotic products and the potential to increase EPS production by co-culture strategies.

Draft genome sequence of Lactobacillus helveticus OSU-BDGOAK2 and Lactobacillus kefiranofaciens OSU-BDGOA1, kefir grain isolates with potential antibacterial activity.

We present the draft genome sequence and assembly of Lactobacillus helveticus OSU-BDGOAK2 and Lactobacillus kefiranofaciens OSU-BDGOA1 isolated from kefir grains that exhibited in vitro antibacterial activity against Escherichia coli ATCC 25922, Listeria innocua ATCC 51742, and Staphylococcus epidermidis ATCC 1222. Genome analysis of both strains revealed gene clusters encoding bacteriocins.

Characteristics of lactose-free/low-lactose fermented dairy products containing microorganisms that are part of the kefir grain consortium: Scoping review

Introduction: There are a significant number of scientific publications dedicated to the analysis of lactose-free and low-lactose fermented milk products, primarily in the context of studying specific microorganisms or starter cultures and their functioning under conditions of limited lactose content. Nevertheless, the synthesis of accumulated data has not been sufficiently highlighted, considering that a large part of the research was conducted over the past 10 years, and an intensive examination of this issue has only begun in the recent past.Purpose: To systematically and critically analyze scientific research that sheds light on the specifics of fermented milk products with a reduced lactose level, focusing on a comparative study of standard fermented milk products and their low-lactose or lactose-free modifications. Special attention is paid to fermented milk products containing microorganisms that are part of the kefir fungus consortium.Materials and Methods: Sources for the review were selected from the Scopus, ScienceDirect, RISC (Russian Index of Science Citation), and Google Scholar databases. Out of 208 publications relevant in terms of keywords, 34 were selected for analysis. The main criterion for selecting articles was a description in them of studies on the use of low-lactose or lactose-free milk as a medium for cultivating starter cultures. This review of the subject area is based on the PRISMA-ScR protocol.Results: Many scientific studies indicate that the prepared low-lactose or lactose-free fermented milk products exhibited similar or improved physicochemical and organoleptic characteristics compared to standard fermented milk products. In experiments aimed at evaluating microbiological characteristics, it was observed that the concentration of lactic acid bacteria in the analyzed products either matched, was lower, or was higher than in traditional fermented milk products. In some publications, scientists found that the final fermented milk product had deteriorated organoleptic and physicochemical parameters. However, all reviewed low-lactose or lactose-free fermented milk products met the standards and regulatory requirements of the countries in which the respective research was conducted.Conclusion: The development of low-lactose and lactose-free fermented milk products is a relevant direction in the food industry, considering their similarity to conventional products and adaptability for consumption by individuals with varying degrees of lactose intolerance. There are nuances related to the duration of the fermentation process, organoleptic characteristics, acidity level, syneresis, preservation, and growth dynamics of microorganisms. These features can be dictated by specific strains of microflora used in the starter and, most often, their influence remains minimal. The results obtained in this study may be useful for selecting starter cultures when designing various fermented milk products, including yogurt, fortified yogurt, kefir, ayran, ryazhenka, and acidophilus. For a deeper understanding of the interaction of the modified glucose-galactose medium with various strains and species of lactic acid bacteria, further experimental work is needed, considering that the activity of these bacteria directly determines the quality characteristics of the final fermented milk product.

Investigation of the water kefir grain formation and growth by image analysis

Investigation of the water kefir grain formation and growth by image analysis

Ulasan Ilmiah: Karakteristik Mutu Nutrisi, Organoleptik dan Mikrobiologis Kefir Susu Kambing

Kefir is a beverage product obtained through the fermentation process of pasteurized milk using a starter from kefir grain. Kefir can be made using goat's milk. Processing of goat milk into kefir is a method used to increase the fat, protein and carbohydrate content of goat milk. There are many factors that affect the quality of kefir. This study aims to determine the potential of of goat's milk kefir based on nutritional, organoleptic and microbiological parameters. The method used is a literature study that comes from various data sources. The selection of cited journals has gone through a screening process based on inclusion, special and exclusion criteria. The results showed, making goat milk kefir requires 12-36 hour of fermentation proces and using a concentration of kefir grains ranging from 2.5 – 5%. This proces, produces a kefir product with good organoleptic characteristics with a white color, distinctive aroma of kefir, sour taste, and soft texture. Goat milk kefir generally has a protein content of 2.96 – 3.66%, fat 2.02 – 5.35%, carbohydrates 2.45 – 5.6%, ash 0.42 – 0.80, ethanol 0.72% with an acidic pH. Goat's milk kefir has a lower lactose concentration of 0.2-0.5% than cow's milk. It also contains various types of microorganisms, including LAB and yeast, which are present in kefir grains which vary widely, with an amount of around 6.4 x 104 to 8.5 x 108 and 1.5 x 105 to 3.7 x 108 cfu/ml. The number of microbes in the kefir has met the standard as a functional food.

Evaluating the roughness dynamics ofkefir biofilms grown onAmazon cupuaçu juice: a monofractal andmultifractal approach.

We conducted a comprehensive analysis of the surface microtexture of kefir biofilms grown on Theobroma grandiflorum Shum (cupuaçu) juice using atomic force microscopy. Our goal was to investigate the unique monofractal and multifractal spatial patterns of these biofilms to complement the existing limited literature. The biofilms were prepared dispersing four different concentrations of kefir grains in cupuaçu juice. Our morphological analysis showed that the surface of the obtained biofilms is essentially formed by the presence of cupuaçu fibers and microorganisms like lactobacilli and yeast. The topographic height-based parameter analysis reveals that there is a dependence between surface roughness and the concentration of kefir grains used. The strongly anisotropic well-centralized topographical height distribution of the biofilms also exhibited a quasi-symmetrical and platykurtic pattern. The biofilms exhibit comparable levels of spatial complexity, surface percolation and surface homogeneity, which can be attributed to their similar topographic uniformity. This aspect was further supported by the presence of similar multifractality in the biofilms, suggesting that despite their varying topographic roughness, their vertical growth dynamics follow a similar pattern. Our findings demonstrate that the surface roughness of kefir biofilms cultivated on cupuaçu juice is influenced by the concentration of kefir grains in the precursor solution. However, this dependence follows a consistent pattern across different concentrations. Graphical Abstract.

A robust nanoLC high-resolution mass spectrometry methodology for the comprehensive profiling of lactic acid bacteria in milk kefir

Consumer attention to functional foods containing probiotics is growing because of their positive effects on human health. Kefir is a fermented milk beverage produced by bacteria and yeasts. Given the emerging kefir market, there is an increasing demand for new methodologies to certify product claims such as colony-forming units/g and bacterial taxa. MALDI-TOF MS proved to be useful for the detection/identification of bacteria in clinical diagnostics and agri-food applications. Recently, LC-MS/MS approaches have also been applied to the identification of proteins and proteotypic peptides of lactic acid bacteria in fermented food matrices.Here, we developed an innovative nanoLC-ESI-MS/MS-based methodology for profiling lactic acid bacteria in commercial and artisanal milk kefir products as well as in kefir grains at the genus, species and subspecies level. The proposed workflow enables the authentication of kefir label claims declaring the presence of probiotic starters. An overview of the composition of lactic acid bacteria was also obtained for unlabelled kefir highlighting, for the first time, the great potential of LC-MS/MS as a sensitive tool to assess the authenticity of fermented foods.

Employing alternative culture media in kefiran exopolysaccharide production: Impact on microbial diversity, physicochemical properties, and bioactivities

Kefiran is a biomaterial with potential application in developing novel materials for food technology. In this study, sugarcane sugar (REF), raw sugar (RAS), brown sugar (BRS), soy molasses (SOM), and sugarcane molasses (SCM) were evaluated for the production of kefiran from kefir biomass rather than cow's milk (CMK), the usual medium. The produced kefiran was purified and characterized by colorimetry, Fourier-transform infrared spectroscopy (FTIR), thermogravimetric analysis, and morphology. It was also assayed for antioxidant and antimicrobial activity. SCM had the highest average daily rate of kefir biomass production (29.17 %/day). The composition of the culture medium affected the microbial diversity of kefir grains, and the Lactobacillus genus was the most abundant (39.8 %, 40.0 %, and 83.9 % to SCM, SOM, and CMK, respectively) in the samples that presented the highest biomass production and kefiran extraction yields. FTIR spectra showed that the bands of kefiran produced in REF and RAS were narrower than those of the other samples. Kefiran grew in an alternative culture medium also exhibited higher thermal stability (Tonset and TMax was higher than 250 and 280 °C, respectively). Those grown in SOM and SCM displayed antimicrobial and antioxidant activities similar to those of kefiran produced in CMK. The results indicate that agro-industrial by-products (SCM and SOM) are potential alternatives for kefiran production from kefir biomass.

Kombucha and Water Kefir Grains Microbiomes' Symbiotic Contribution to Postbiotics Enhancement.

Wild artisanal cultures, such as a symbiotic culture of bacteria and yeasts (SCOBY) and water kefir grains (WKG), represent a complex microorganism consortia that is composed of yeasts and lactic and acetic acid bacteria, with large strains of diversity and abundance. The fermented products (FPs) obtained by the microbiome's contribution can be included in functional products due to their meta-biotics (pre-, pro-, post-, and paraprobiotics) as a result of complex and synergistic associations as well as due to the metabolic functionality. In this study, consortia of both SCOBY and WKG were involved in the co-fermentation of a newly formulated substrate that was further analysed, aiming at increasing the postbiotic composition of the FPs. Plackett-Burman (PBD) and Response Surface Methodology (RSM) techniques were employed for the experimental designs to select and optimise several parameters that have an influence on the lyophilised starter cultures of SCOBY and WKG activity as a multiple inoculum. Tea concentration (1-3%), sugar concentration (5-10%), raisins concentration (3-6%), SCOBY lyophilised culture concentration (0.2-0.5%), WKG lyophilised culture concentration (0.2-0.5%), and fermentation time (5-7 days) were considered the independent variables for mathematical analysis and fermentation conditions' optimisation. Antimicrobial activity against Bacillus subtilis MIUG B1, Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Aspergillus niger MIUG M5, antioxidant capacity (DPPH), pH and the total acidity (TA) were evaluated as responses. The rich postbiotic bioactive composition of the FP obtained in optimised biotechnological conditions highlighted the usefulness of the artisanal co-cultures, through their symbiotic metabolic interactions for the improvement of bioactive potential.

Cover Caption

AbstractJuly Online Cover: Relative abundance of high‐quality reads at species levels of bacteria and fungi in the metagenome of milk kefir and kefir grains, and taxonomic profile of kefir metagenomes, from “Metagenomic analysis and antibacterial activity of kefir microorganisms” by Brianda D. González‐Orozco, Israel García‐Cano, Alejandra Escobar‐Zepeda, Rafael Jiménez‐Flores, and Valente B. Álvarez. p. 2936–2952.

Evaluation of mid-infrared spectra associated with chemometrics for the determination of physicochemical properties during fermentation of a new strawberry-based beverage with water kefir grains

This study aimed to explore the mid-infrared technique for the characterization of a non-dairy fermented beverage. Fruit juice was used as a substrate for fermentation made by microorganisms from water kefir. The fermented beverage was evaluated by standardized methods regarding pH, total titratable acidity, soluble solids, and total reducing sugars. Then, infrared spectra were obtained and calibration models were built to predict these parameters in the fermented beverage. All obtained models showed high coefficients of determination values (R2) in the calibration and prediction stages, in addition to high residual predictive deviation (RPD) values. For the prediction data, the values of R² were equal to 0.91, 0.93, 0.95, and 0.91 for pH, total titratable acidity, soluble solids, and total reducing sugars, respectively. It was evidenced that the soluble solids content was the most suitable physicochemical property for estimation by the elaborated calibration models (higher R2 and RPD values). The results showed that such parameters could be estimated in kefir quickly through the mid-infrared analysis.

Effects of High-Intensity Ultrasound Pretreatment on the Exopolysaccharide Concentration and Biomass Increase in Cheese Whey Kefir

Cheese whey (CW) is the liquid by-product of cheese and yogurt making. This potential pollutant has high-quality nutrients exploitable through fermentation processes. Using high-intensity ultrasound on dairy products has shown several technological advantages for bioprocesses. Therefore, this study aimed to investigate the effect of high-intensity ultrasound (HIUS) on kefir grains biomass increase and specific metabolites in CW kefir. Fresh CW was ultrasonicated at 9.0 ± 2.7 and 18.0 ± 3.0 W/cm2 for 30 and 180 s, inoculated with kefir grains, and fermented for 40 h. Total exopolysaccharide production, kefir grains biomass increase, titratable acidity, pH, and soluble solids were analyzed every 8 h. CW pretreated with 18.0 ± 3.0 W/cm2 for 180 s and fermented for 16 h had significantly higher (p < 0.05) total exopolysaccharide concentration than the control: 212.7 ± 0.0 and 186.6 ± 0.0 mg/L, respectively. Ultrasonicated CW at 18 W/cm2 for 30 and 180 s at 24 h fermentation time had significantly higher kefir grains biomass (p < 0.05) than the control: 44.2 ± 0.8 and 43.6 ± 0.9 g/L, and 40.5 ± 0.4 g/L, respectively. Fresh CW pretreated with HIUS enhanced the biosynthesis of kefir beverage total exopolysaccharides concentration and kefir grains biomass.

Microbial and Chemical Changes during Fermentation of Coconut Water Kefir Beverage

This study investigated changes in the microbial growth and chemical characteristics of coconut water kefir (CWK) during fermentation. The carbohydrate profile, in terms of glucose and sucrose consumption, production of carboxylic acids, and changes in amino acid profile, was determined during CWK fermentation over a period of 96 h. The results showed that the kefir grains were able to utilise both glucose and sucrose and produce significant quantities of carboxylic acids after 96 h of fermentation. The total titratable acidity significantly increased throughout 96 h of incubation at 30 °C, which correlated to a significant drop in pH to 2.8 for CWK supplemented with 12 g/L of sucrose. In addition, this was accompanied by a significant increase in lactic acid, acetic acid, and pyruvic acid. During fermentation, a total of eighteen amino acids were generated, with a notable decline observed across all amino acids. Among them, glutamic acid exhibited a higher concentration compared to the other amino acids. The Scanning Electron Microscope (SEM) results confirmed a higher density of lactic acid bacteria (LAB) and acetic acid bacteria (AAB), with fewer yeast cells through morphological identification. Overall, the findings support the notion that coconut water fermented with kefir could be used as a potential functional starter to produce other fermented food products or a refreshing beverage.

Effect of Agave Fructans on Changes in Chemistry, Morphology and Composition in the Biomass Growth of Milk Kefir Grains.

Prebiotic effects have been attributed to agave fructans through bacterial and yeast fermentations, but there are few reports on their use as raw materials of a carbon source. Kefir milk is a fermented drink with lactic acid bacteria and yeast that coexist in a symbiotic association. During fermentation, these microorganisms mainly consume lactose and produce a polymeric matrix called kefiran, which is an exopolysaccharide composed mainly of water-soluble glucogalactan, suitable for the development of bio-degradable films. Using the biomass of microorganisms and proteins together can be a sustainable and innovative source of biopolymers. In this investigation, the effects of lactose-free milk as a culture medium and the addition of other carbon sources (dextrose, fructose, galactose, lactose, inulin and fructans) in concentrations of 2, 4 and 6% w/w, coupled with initial parameters such as temperature (20, 25 and 30 °C), % of starter inoculum (2, 5 and 10% w/w) was evaluated. The method of response surface analysis was performed to determine the optimum biomass production conditions at the start of the experiment. The response surface method showed that a 2% inoculum and a temperature of 25 °C were the best parameters for fermentation. The addition of 6% w/w agave fructans in the culture medium favored the growth of biomass (75.94%) with respect to the lactose-free culture medium. An increase in fat (3.76%), ash (5.57%) and protein (7.12%) content was observed when adding agave fructans. There was an important change in the diversity of microorganisms with an absence of lactose. These compounds have the potential to be used as a carbon source in a medium culture to increase kefir granule biomass. There was an important change in the diversity of microorganisms with an absence of lactose, where the applied image digital analysis led to the identification of the morphological changes in the kefir granules through modification of the profile of such microorganisms.

Multi-Component Biodegradable Materials Based on Water Kefir Grains and Yeast Biomasses: Effect of the Mixing Ratio on the Properties of the Films.

The use of biopolymeric materials is restricted for some applications due to their deficient properties in comparison to synthetic polymers. Blending different biopolymers is an alternative approach to overcome these limitations. In this study, we developed new biopolymeric blend materials based on the entire biomasses of water kefir grains and yeast. Film-forming dispersions with varying ratios of water kefir to yeast (100/0, 75/25, 50/50 25/75 and 0/100) underwent ultrasonic homogenisation and thermal treatment, resulting in homogeneous dispersions with pseudoplastic behaviour and interaction between both biomasses. Films obtained by casting had a continuous microstructure without cracks or phase separation. Infrared spectroscopy revealed the interaction between the blend components, leading to a homogeneous matrix. As the water kefir content in the film increased, transparency, thermal stability, glass transition temperature and elongation at break also increased. The thermogravimetric analyses and the mechanical tests showed that the combination of water kefir and yeast biomasses resulted in stronger interpolymeric interactions compared to single biomass films. The ratio of the components did not drastically alter hydration and water transport. Our results revealed that blending water kefir grains and yeast biomasses enhanced thermal and mechanical properties. These studies provided evidence that the developed materials are suitable candidates for food packaging applications.

Iranian traditional kefir beverage: isolation and identification of beneficial microorganisms and evaluation of antimicrobial activity against food-borne pathogens

Purpose Kefir is a traditional fermented dairy beverage that has numerous health benefits due to the presence of bacteria and yeasts in an exopolysaccharide matrix. This study aims to isolate and identify beneficial microorganisms and evaluate the antimicrobial activity of kefir beverage against two important food-borne pathogens including Salmonella Typhimurium and Listeria monocytogenes. Design/methodology/approach Microorganisms were identified by the polymerase chain reaction with specific primers, and antimicrobial activity was evaluated by the disk diffusion method. Findings The following microorganisms were identified as natural inhabitants of the kefir grains: Leuconostoc lactis, Lactococcus lactis subspecies lactis, Streptococcus cremoris, Enterococcus faecalis, Enterococcus faecium, Lactobacillus helveticus, Leuconostoc mesenteroides, Lactobacillus fermentum, Lactobacillus reuteri, Lactobacillus plantarum, Lactobacillus casei, Bifidobacterium langum, Saccharomyces cerevisiae and Pichia fermentas. Also, the results obtained from the disk diffusion method showed the inhibitory effect of kefir milk on Salmonella Typhimurium and Listeria monocytogenes with an inhibitory average diameter of 8.3 ± 4–9.1 ± 2.8 and 3.4 ± 3–6.6 ± 3 mm, respectively. Originality/value The results of this study showed that Iranian traditional kefir beverage contained different species of lactic acid bacteria and yeasts and has antimicrobial activity against two important food-borne pathogens, Salmonella Typhimurium and Listeria monocytogenes, which the highest inhibitory effect was observed against Salmonella Typhimurium.

Lactic Acid Bacteria and Yeasts from Milk Kefir Grains: Isolation, Characterization, Screening, and Identification

Abstract Lactic acid bacteria and yeast are abundant in kefir grains. Six natural kefir grains were used in this research to isolate and purify 64 strains of yeast and 108 strains of lactic acid bacteria. A total of three lactic acid bacteria and one yeast strain (Lactococcus lactis 2C6, Lactobacillus helveticus 6171, Lactobacillus plantarum 4M2 and Saccharomyces cerevisiae 6Y6) were examined for their ability to protein hydrolysis, water holding capacity, alcohol production, acid and bile salt tolerance and identified by 16S rDNA. The proteolytic ability, alcohol generation, and acid and bile salt tolerance of 4M2 and 6Y6 were excellent. Lyophilization was used to create bacterial powder, which laid the foundation for the subsequent development of direct-vat-set (DVS) kefir starter.

Microbiological profiles, physiochemical properties and volatile compounds of goat milk kefir fermented by reconstituted kefir grains

Kefir is a health-promoting fermented dairy products, consisting in a variety of bacterial and fungal species. The physical and chemical properties and the correlation between the microbiota and volatile compounds of goat milk kefir fermented by reconstituted kefir grains were investigated. Cluster analysis showed that a strong symbiotic homeostasis system existed in goat milk kefir, indicating that the composition of microorganism in reconstituted kefir grains was still restricted by the original one. Compared with other kefir samples, goat milk kefir 80DE20CN (ratios of kefir grains CN/DE = 80/20) showed significant improvement in physiochemical properties and “Fruity” flavor. Positive correlations among Lactobacillus helveticus and ethyl acetate, amyl acetate and ethanol, Kazachstania unispora and octyl formate, 1-heptanol and hexanoic acid were obtained by Partial least squares regression analysis (PLSR) model. The results showed that changes in microbial succession patterns may be useful to optimize the physiochemistry and flavor of goat milk kefir or related products.

Copper-Contaminated Substrate Biosorption by Penicillium sp. Isolated from Kefir Grains.

In this bioremediation study, the fungus Penicillium sp. isolated from kefir grains was evaluated for its resistance to copper in the culture medium. Penicillium sp. was cultivated in liquid medium prepared using 2% malt-agar at pH 7.0. Biomass of the fungus was significantly reduced, but only when 800 mg·L-1 of Cu(NO3)2 copper nitrate was used. The effect on radial growth of the fungus in experiments combining different pH values and the inorganic contaminant showed an inhibition of 73% at pH 4.0, 75% at pH 7.0 and 77% at pH 9.0 in liquid medium. Thus, even though the growth of Penicillium sp. could be inhibited with relatively high doses of copper nitrate, images obtained with scanning electron microscopy showed the preservation of fungal cell integrity. Therefore, it can be concluded that Penicillium sp. isolated from kefir grains can survive while performing bioremediation to minimize the negative effects of copper on the environment through biosorption.