Cheese Varieties Research Articles

A brief identification of lactic acid bacteria isolated from Algerian goat’s Jben by MALDI-TOF MS

In this paper, we investigated the lactic acid bacteria’s community of the Algerian goat’s Jben in order to define and preserve it. This cheese variety is only handmade with raw milk, and a dried flower of Cynara cardunculus is used instead of the animal rennet, it is also restricted in sub-Saharan prov-inces in Algeria, and no previous studies on its lactic acid bacteria composi-tion have been performed before. Matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) procedure was used to identify 36 lactic acid bacteria isolated from artisanal Jben made from Arabia goat’s raw milk in Naâma (Algeria). The results showed the domination of the Enterococcus genus presents with four species: E. durans, E. faecalis, E. hirae and E. faecium. Lactococcus lactis and Leuconostoc mesenteroïdes were also detected. The species identified were favoured by the composition of goat’s raw milk microflora used and the artisanal manufacturing process of the cheese. The use of MALDI-TOF MS method provided us with a rapid diagnosis of the Jben lactic acid bacteria’s community. This method based on microbial abundant proteins fingerprint diagnosis was chosen for its fast, precise, low cost and less complicated analyse.

Non-canonical Activities of Hog1 Control Sensitivity of Candida albicans to Killer Toxins From Debaryomyces hansenii.

Certain yeasts secrete peptides known as killer toxins or mycocins with a deleterious effect on sensitive yeasts or filamentous fungi, a common phenomenon in environmental species. In a recent work, different Debaryomyces hansenii (Dh) strains isolated from a wide variety of cheeses were identified as producing killer toxins active against Candida albicans and Candida tropicalis. We have analyzed the killer activity of these toxins in C. albicans mutants defective in MAPK signaling pathways and found that the lack of the MAPK Hog1 (but not Cek1 or Mkc1) renders cells hypersensitive to Dh mycocins while mutants lacking other upstream elements of the pathway behave as the wild type strain. Point mutations in the phosphorylation site (T174A-176F) or in the kinase domain (K52R) of HOG1 gene showed that both activities were relevant for the survival of C. albicans to Dh killer toxins. Moreover, Hog1 phosphorylation was also required to sense and adapt to osmotic and oxidative stress while the kinase activity was somehow dispensable. Although the addition of supernatant from the killer toxin- producing D. hansenii 242 strain (Dh-242) induced a slight intracellular increase in Reactive Oxygen Species (ROS), overexpression of cytosolic catalase did not protect C. albicans against this mycocin. This supernatant induced an increase in intracellular glycerol concentration suggesting that this toxin triggers an osmotic stress. We also provide evidence of a correlation between sensitivity to Dh-242 killer toxin and resistance to Congo red, suggesting cell wall specific alterations in sensitive strains.

Cardoon-based rennets for cheese production.

The use of crude aqueous extracts of Cynara cardunculus flowers as coagulants in the production of high-quality sheep and goat cheeses-as are the cases of several Portuguese and Spanish cheese varieties with Protected Designation of Origin status-has been maintained since ancient times. The unique rheological attributes and sensory properties characteristic of these cheeses have always suggested that this plant coagulant (and, therefore, its isolated milk-clotting proteases) could be used as alternative rennet in the dairy industry, particularly suited for the production of sheep and goat cheeses. However, the lack of standardization of C. cardunculus crude flower extracts, whose quality and performance depends on numerous factors, has always hampered the application of this plant rennet in industrial production scales. To overcome these limitations, and to aim at developing more effective solutions with potential for scalability of production and commercial application, several strategies have been undertaken in more recent years to establish new cardoon-based rennets. This review provides an overview on these developments and on the currently available solutions, which range from producing standardized formulations of native cardoon enzymes, to the optimization of the heterologous production of cardosins and cyprosins to generate synthetic versions of these milk-clotting enzymes. Challenges and emerging opportunities are also discussed.

Evidence of low levels of aflatoxin M1 in milk and dairy products marketed in Qatar

This study was designed to explore the incidence of aflatoxin M1 (AFM1) in milk and dairy products marketed in Qatar. Milk (n = 72), yogurt (n = 21), cheese (n = 46), butter (n = 18) and laban (n = 25) samples were initially screened by competitive enzyme linked immunosorbent assay (ELISA) for the presence of AFM1, followed by confirmation with ultra-high-performance liquid chromatography (UHPLC). AFM1 was detected in 85%, 76%, 85%, 67% and 76% of the milk, yogurt, cheese, butter and laban samples, respectively. The levels of AFM1 in pasteurized vs ultra-high temperature (UHT) milk and full-vs low-fat yogurts showed a non-significant difference. Halloumi and Kashkaval cheese varieties showed a significantly higher AFM1 contents as compared to Mozzarella, Edam, Cheddar, cream and Moshalal cheese. Likewise, unsalted butter demonstrated significantly (p < 0.05) higher AFM1 values than salted butter samples. None of the tested samples presented AFM1 levels above the EU maximum limits of 50 ng/L or kg for milk, yogurt and butter, and 250 ng/kg for cheese. From the finding of present study, it can be concluded that, although high percentage of dairy products marketed in Qatar demonstrated AFM1 contents, but do not represent a public health concern considering the EU maximum limits.

Chemical composition and nutritional quality of short-ripened rennet cheeses produced by traditional methods

The aim of the study was to determine the variability in the chemical composition and nutritive value parameters of smoked and unsmoked short-ripened rennet cheeses and unsmoked long-ripened rennet cheeses produced by traditional methods. The raw material for the production of short-ripened cheeses was pasteurized cows’ milk obtained from a dairy, whereas the long-ripened cheeses were manufactured from raw cows’ milk obtained from the producer’s farm. All three varieties of cheese examined were produced with commercial dairy starter cultures. The material for the study was collected in winter, directly at the producers’ retail outlets in southern and eastern Poland. The basic chemical composition was determined according to the Polish Standards, whereas the amino acid profiles of proteins from the cheeses were determined by ion-exchange chromatography. The result analysis revealed significant differences between the different varieties of cheese in terms of their water content, ranging from 32.2% to 42.1%, as well as protein content, which varied from 25.6% to 31.6%. Fat levels ranged between 22.2% and 24%, whereas total ash content amounted to 5.1-5.8%. The significantly highest salt content was found in unsmoked short-ripened cheeses. In all three cheese varieties, the total exogenous amino acid content was comparable, ranging from 46.17 g to 47.36 g/100 g protein, and that of endogenous amino acids varied from 52 g to 53 g/100 g protein. The biological value of proteins was determined by calculating to the chemical score (CS), as described by Mitchell and Block, and the essential amino acid index (EAAI), as described by Oser. A comparison of the results with the standard chicken egg white proteins showed that the limiting amino acids for all varieties of cheese were methionine and cysteine. On the other hand, a comparison with the FAO/WHO-suggested pattern of amino acid requirements (1991) for all age groups over 1 year of age showed that the limiting amino acids were methionine and cysteine in smoked and unsmoked short-ripened cheeses, and treonine in long-ripened cheeses. Considering the chemical indices, such as CS and EAAI, it may be concluded that the traditional rennet cheeses produced in southern and eastern Poland have a favourable amino acid composition of proteins and a high nutritive value..

Long-Term Ripening Evaluation of Ewes’ Cheeses by Fourier-Transformed Infrared Spectroscopy under Real Industrial Conditions

The ripening changes over time of special cheeses (Pecorino, ewes’ ripe, and Gouda) made with ewes’ milk were evaluated using FTIR/ATR spectroscopy during approximately one year. The midinfrared FTIR/ATR analyses were carried out in different ripening times between the cheese varieties and processed by means of multivariate statistical approaches. Overall, during the maturation, we observed a downward trend of the absorbance intensity of the amide group peaks (1700 to 1500 cm−1), which is linked to the breakdown of peptide bonds. Similar behavior was obtained for the lipidic region (3000 to 2800 cm−1 and 1765 to 1730 cm−1). Hierarchical cluster analysis and principal component analysis allowed the evaluation of the physicochemical changes of the cheeses. The proteolysis occurs in a fast pace during the first trimester of the ripening process, and the lipids are converted to smaller species as the times goes by. Our results indicate that infrared spectroscopy can be a useful tool in determining optimal temporal parameters in stages involving the development, production, and even a possible estimation of shelf life of cheeses.

Application of multiple factor analysis for the descriptive sensory evaluation and instrumental measurements of bryndza cheese as affected by vacuum packaging

ABSTRACTMultiple factor analysis was used to examine naturally white, mature and spreadable bryndza cheese varieties. Sensory and physiochemical properties were evaluated during storage in vacuum and non-vacuum packed conditions at 4 ± 2°C. The results of statistical method showed similarity in varieties made from pasteurized ewe milk and mixed bryndza cheese before and after storage at both package conditions (p < 0.05). The statistical method used in the study was a suitable tool for assessing differences in food quality and provided an opportunity to visualize and categorize the results with regard to shelf life and packaging.

Traditional Italian cheeses: Trace element levels and estimation of dietary intake

Concentrations of some toxic (Hg, Cd, Pb) and essential metals (Zn, Cu, Cr, Fe) relevant to human health were determined in different varieties of traditional cheeses, included in Traditional Agricultural Food Products (TAP) list, sampled in various farms in Apulia and Sicily, two regions of Southern Italy. Among toxic metals, Pb showed the highest concentrations (hard cheese: 0.13 μg g−1 w.w.; fresh cheese: 0.07 μg g−1 w.w.), while among essential element the most abundant was Fe (hard cheese: 2.86 μg g−1 w.w.; fresh cheese: 2.23 μg g−1 w.w.). In terms of food safety, the toxic metal concentrations in the majority of the samples investigated were below the maximum permitted levels set by several non-European countries, except for Pb showing levels notably exceeding in some hard cheese (0.22–0.37 μg g−1 w.w.). Also the estimated intakes of toxic metals remained far below the provisional tolerable intakes and consequently no health risk is expected from the consumption of these cheeses. However, for greater food safety, besides to a continuous surveillance it would be advisable to establish maximum limits for metals either in milk or products made of milk.

Handling, Processing, Utilization and Marketing of &amp;lt;i&amp;gt;Ayib&amp;lt;/i&amp;gt; (Ethiopian Traditional Cottage Cheese) Varieties Produced in Selected Areas of Eastern Gojjam, Northwestern Highlands of Ethiopia

The study was conducted to asses handling, processing, utilization, marketing as well as other socio-economic benefits of traditional cheese varieties making and to identify major constraints of traditional cheese varieties production in Awabal, Gozamin, Machakel and Dejen districts of Eastern Gojjam. A cross-sectional survey was conducted to collect data. A total of 240 smallholder dairy producers (60 households from each district) were individually interviewed using a semi-structured questionnaire. The major traditional cheese varieties produced in the study areas include Ayib, Metata, Hazo and Zureshekefign. Among these, three of them (Ayib, Metata and Hazo), are dominant. Respondents indicated that Metata cheese variety has high nutritional values and cure seven types of disease namely malaria and yellow fever, depressed appetite; Ejeseb (stomach upset), metabolic disorder (bloating), gastro-intestinal parasites, prevent nausea and digestion upset. The respondents reported twenty kinds of spices blended at one time during Metata cheese preparation and five kinds of spices commonly added into Ayib during Ayib making. Various types of herbs and plant species are used for smoking and washing of utensils during production of traditional cheese varieties in the study areas. The major marketing constraints of traditional cheese varieties were absence of sufficient buyer in the market, traditional taboos to sell cheese varieties in an open market, seasonal demand and supply, distance to the market, transportation access and quality. The major socio-economic benefits of traditional cheese varieties were to fulfill household necessities, sources of additional income, to purchase dairy inputs and for children school fee. Further scientific study needs to be conducted in order to confirm nutritional and medicinal properties of Metata cheese variety. Therefore the bio-active components of these spices and their effect on shelf life, nutritional value, medicinal uses and other attributes on traditional cheese varieties needed further investigation.

Environmental Awareness of University Students on White Cheese Waste Water

Universal environmental contamination is a real situation that deteriorates our world step by step. The dairy factory out flowing is the second greatest source of contamination in water streams. The environmental impact of these factories can be very high, especially due to the discharge of wastewater with high content of organic matter and nutrients. These problems can be analyzed only after performing factual study of various physicochemical characteristics. In the presented study, physicochemical parameters like, temperature, pH, COD, TDS, TS and SS were taken into account. Another purpose of this study is to ascertain university student’s awareness and consciousness against general and dairy products related environments. These levels were evaluated by the survey method. The data obtained from the questionnaire were analyzed using SPSS 20.0. Results showed that the public and company owners/ employees in particular should be informed about the seasonal and cheese variety dependent patterns in environmental pollution.

The Effect of Different Methods of Fermentation on the Detection of Milk Protein Residues in Retail Cheese by Enzyme‐Linked Immunosorbent Assay (ELISA)

Many commercially available enzyme-linked immunosorbent assays (ELISAs) are not validated for detection of allergenic residues in fermented or hydrolyzed products. This research seeks to determine if commercial milk ELISAs can detect milk residues in varieties of cheese that have undergone different styles of fermentation and different degrees of proteolysis. Only certain milk ELISA kits are capable of detecting residues in all varieties of cheese. However, commercial milk ELISA kits are capable of semiquantitative detection of cheese residues in foods, or in industry settings for the validation of allergen cleaning programs.

Effects of pre-cooked cheeses of different emulsifying conditions on mechanical properties and microstructure of processed cheese

The effect of pre-cooked cheeses of different emulsifying conditions on the viscosities, mechanical properties, fat globules, and microstructure of processed cheese was investigated, and changes in protein network relating to the creaming effect and the occurrence of yielding point were discussed. The addition of pre-cooked cheeses with a short stirring time had no obvious impact on the fat globules and protein network. The random network brought low viscosities and a gradual increase in the fracture stress/strain curve. The addition of pre-cooked cheeses with the long stirring time caused protein network to become fine-stranded. The fine-stranded network caused creaming effect, and brought yielding points in the mechanical properties. The pre-cooked cheese with the small fat globules also caused fat globules to become smaller, and give the processed cheese more firmness. This study provides a potential solution to control the functional properties of processed cheese by using a variety of pre-cooked cheeses.

Direct analysis of volatile organic compounds in foods by headspace extraction atmospheric pressure chemical ionisation mass spectrometry.

RationaleThe rapid screening of volatile organic compounds (VOCs) by direct analysis has potential applications in the areas of food and flavour science. Currently, the technique of choice for VOC analysis is gas chromatography/mass spectrometry (GC/MS). However, the long chromatographic run times and elaborate sample preparation associated with this technique have led a movement towards direct analysis techniques, such as selected ion flow tube mass spectrometry (SIFT‐MS), proton transfer reaction mass spectrometry (PTR‐MS) and electronic noses. The work presented here describes the design and construction of a Venturi jet‐pump‐based modification for a compact mass spectrometer which enables the direct introduction of volatiles for qualitative and quantitative analysis.MethodsVolatile organic compounds were extracted from the headspace of heated vials into the atmospheric pressure chemical ionization source of a quadrupole mass spectrometer using a Venturi pump. Samples were analysed directly with no prior sample preparation. Principal component analysis (PCA) was used to differentiate between different classes of samples.ResultsThe interface is shown to be able to routinely detect problem analytes such as fatty acids and biogenic amines without the requirement of a derivatisation step, and is shown to be able to discriminate between four different varieties of cheese with good intra and inter‐day reproducibility using an unsupervised PCA model. Quantitative analysis is demonstrated using indole standards with limits of detection and quantification of 0.395 μg/mL and 1.316 μg/mL, respectively.ConclusionsThe described methodology can routinely detect highly reactive analytes such as volatile fatty acids and diamines without the need for a derivatisation step or lengthy chromatographic separations. The capability of the system was demonstrated by discriminating between different varieties of cheese and monitoring the spoilage of meats.

Measuring antibiotic resistance in mixed cultures: Isothermal microcalorimetry as a novel analytical tool

Most Swiss cheese varieties use raw, undefined mixed cultures, which are composed of strains of Lactobacillus delbrueckii subsp. lactis and Streptococcus thermophilus. Single strains of these consortia are not always available. The aim of this study was to determine the antibiotic resistance of such consortia and propose a new approach to assess the antibiotic resistance. In this work, we developed a new method using isothermal microcalorimetry to assess antibiotic resistance in raw mixed cultures. For most of these cultures, no transferable antibiotic resistance genes were detected. In most cases, the results showed that all raw mixed cultures have lower minimal inhibitory concentration (MIC) than the European Food Safety Authority-defined levels, with the exception of the aminoglycosides kanamycin and streptomycin where the MIC was >64 mg L−1 for each raw mixed culture.

Effect of high-pressure treatments on proteolysis, volatile compounds, texture, colour, and sensory characteristics of semi-hard raw ewe milk cheese

High pressure (HP) offers potential industrial applications in cheese preservation, but it is essential to provide knowledge concerning their effects on the ripening process and sensory characteristics. In this study, we investigated the effect of different HP treatments (200–500MPa at 14°C for 10min on day 7) on proteolysis, texture, colour, volatile compounds and sensory characteristics of semi-hard raw ewe milk cheese. HP treatments did not affect pH or dry matter values of 60-day-old cheeses. Treatments at pressure levels up to 400MPa led to significant (P<0.01) increases in the total free amino acids (FAA) content at 60days, compared to control cheese, although the cell-free aminopeptidase activity was lower. HP retarded the formation of some volatile compounds in cheeses, the number of compounds affected by HP being higher as the pressure level increased. Cheeses pressurized at 300–500MPa had lower levels of 2-butanone, 2-butanol, 2-propen-1-ol, 1-butanol and acetic acid than control cheese, cheeses pressurized at 400–500MPa lower levels of 1-propanol, 2-pentanol, and butyric and hexanoic acids, and cheeses pressurized at 500MPa lower levels of ethanol, 3-methyl-1-butanol and 3-methyl-2-buten-1-ol. All HP-treated cheeses showed higher fracturability values, and higher Hue angle and lower a* values than control cheese. Despite the differences detected by instrumental analyses between HP-cheeses and control cheese, few significant differences were found between the sensory characteristics of HP-cheeses and control cheese. Only the pressurization of cheese at 500MPa affected significantly (P<0.01) some of the sensory characteristics, with a negative effect on taste intensity but a positive effect on aroma quality.In summary, HP treatments at 200 and 300MPa showed the mildest effects on the characteristics of semi-hard raw ewe milk cheese. HP treatment of this cheese variety at 300, 400 and 500MPa prevented late blowing defect caused by Clostridium tyrobutyricum (Ávila et al., 2016, Food Microbiol. 60, 165–173). Thus, it may be concluded that HP treatment at 300MPa is the most adequate procedure, able to prevent late blowing with minimum changes in cheese characteristics.

Utilization of Edible Films and Coatings as Packaging Materials for Preservation of Cheeses

The diversity of cheese varieties, dynamic microbiological and biochemical changes which undergo during cheese manufacture, ripening and marketing need special care to meet the desired consumer acceptability and satisfaction. In this area packaging plays an important role in cheese industry. Application of films or coatings to cover cheese surfaces has long been recognized as an important treatment for the protection of the quality and safety of the product. The growing environmental concern about the use of un-biodegradable materials in cheese packaging encouraged research to develop edible materials for coating and packaging of cheese. Several polysaccharides and proteins with or without plasticizers, antimicrobial agents and nanoparticles have been used as basis for the development of edible cheese coating. The present manuscript gives an overview on the development and use of edible cheese coating in relation to the cheese quality and safety.

Effect of enzymatic transglutaminase treatment on textural and sensory properties of low-fat UF-Feta cheese incorporated with whey proteins using response surface optimization.

شواهد علمی نشان داده است که مصرف چربی ارتباط مستقیمی با بیماری های مختلفی از جمله چاقی، دیابت، سفت شدن دیواره رگ ها و فشار خون دارد. به همین دلیل طی سال های اخیر تقاضا برای محصولات کم‌چرب افزایش چشمگیری پیدا کرده است. پنیر به‌عنوان یک فراورده لبنی با ارزش نقش پررنگی در رژیم غذایی جامعه دارد. با این حال این فراورده حاوی مقادیر بالایی از چربی می باشد که سبب ایجاد نگرانی های از جانب مصرف‌کنندگان شده است. در این پژوهش از روش سطح پاسخ به‌منظور مطالعه اثر آنزیم ترانس گلوتامیناز (2-0 واحد به ازای هرگرم پروتئین)، محلول کنسانتره پروتئینی آب پنیر یا WPC (16-0 درصد) و سطوح چربی (10-0 درصد) بر خواص کیفی و بهینه‌سازی فرمولاسیون پنیر فراپالایش کم‌چرب استفاده گردید. نتایج نشان داد که کاهش چربی باعث افزایش معنی‌دار سفتی، ارتجاع‌پذیری و میزان رطوبت می شود اما چسبندگی و امتیاز پذیرش کلی نمونه های پنیر کاهش می یابد. برخلاف WPC، تیمار آنزیمی با ترانس گلوتامیناز سفتی، پیوستگی و ارتجاع‌پذیری را بطور معنی‌داری افزایش داد اما میزان رطوبت و چسبندگی پنیر را کاهش داد. در این میان آنزیم ترانس گلوتامیناز اثر معنی‌داری بر امتیاز کلی ارزیابی حسی نداشت اما خواص حسی نمونه‌های پنیر کم‌چرب با جایگزینی WPC با ناتراوه تا میزان 8% بهبود یافت. نتایج بهینه‌سازی با طرح مرکب مرکزی نشان داد که بهترین نمونه با خواص حسی و بافتی مطلوب زمانی حاصل می‌شود که فرمول پنیر شامل 95/5% چربی، 56/0 واحد آنزیم ترانس گلوتامیناز و 79/8% محلول WPC باشد.

Use of smear bacteria and yeasts to modify flavour and appearance of Cheddar cheese

The strains Staphylococcus saprophyticus DPC5671 and Corynebacterium casei DPC5298 were applied in combination with Debaryomyces hansenii DPC6258 to the surface of young Cheddar cheese curd to obtain two different smear-ripened cheeses. A surface microbiota developed over the incubation period, comprising of both yeast and bacteria; pulsed field gel electrophoresis confirmed that the inoculated strains of S. saprophyticus DPC5671 or C. casei DPC5298 were the dominant bacterial strains on the surface of the cheese at the end of the ripening period. The smear cultures changed the appearance and aroma, which were significantly different from the control cheese. The approach presented in this study represents a method for the development of new cheese varieties with novel aromas within a short ripening time.

Lipolysis profile of some non-European raw milk cheese varieties: a review

&lt;p&gt;This review focuses on the lipolysis profile of some non European raw milk cheese varieties. Lipolysis data of some traditional/artisanal non-European raw milk cheeses such as cheeses ripened in an animal skin, cheeses ripened in brine, cheeses produced in small scale by local dairies etc., are collected and presented in this study. The main characteristics of these cheeses and their lipolysis profile in general are discussed. Factors such as the type of cheese, the technology applied for cheese making and the age of cheese are shown to influence the lipolysis profile.&lt;/p&gt;

Bacterial ecology of artisanal Minas cheeses assessed by culture-dependent and -independent methods

Artisanal Minas cheese is produced in Minas Gerais state, Brazil and its varieties are named according to their geographical origin (Serro, Canastra, Serra do Salitre, Araxá and Campo das Vertentes). The cheese is produced with raw cow's milk and the whey from the previous cheese production (“pingo”). The high economic and cultural importance of artisanal cheese in Brazil justifies the efforts to ensure its safety, quality and provenance. This study aimed to characterize the microbial diversity composition, and geographical distribution of artisanal Minas cheese, focusing on the characterization of its autochthonous lactic acid bacteria (LAB) microbiota. Artisanal Minas cheese varieties from Serro, Canastra, Serra do Salitre, Araxá and Campo das Vertentes were analyzed by culture-dependent (culturing and LAB sequencing) and -independent (repetitive extragenic palindromic-PCR (rep-PCR) and length heterogeneity-PCR, LH-PCR) methods to characterize the microbiota. The microbial counts were variable between cheese samples, and some samples presented high number of coagulase positive bacteria and coliforms that may be associated with hygienic issues. In all samples was observed a prevalence of LAB. 16S rRNA sequencing and rep-PCR of the LAB strains identified four genus (Lactobacillus, Lactococcus, Enterococcus and Weissella), ten species and more than one strain per species. Lactobacillus was the most prevalent genera in all the cheeses. LH-PCR revealed a further six genera and ten species that were not identified by culturing, highlighting the importance of combining both culture-dependent and -independent methods to fully characterize microbiota diversity. Principal component analysis of the LH-PCR data and cluster analysis of rep-PCR data revealed that the artisanal Minas cheese microbiota was influenced not only by their geographical origin but also by the cheese farm. The lack of standardization in the milking and cheese manufacturing procedures between artisanal cheese farms could explain the microbial diversity.