Introduction

The incidence of hypercholesterolemia is increasing swiftly with the improvements of people’s living standards and amendments in lifestyles. Elevated serum cholesterol level is usually considered to be the most imperative risk factor for the development of cardiovascular diseases (CVDs), such as atherosclerosis, hypertension, coronary heart disease, and stroke. Both drug therapy and non-pharmacological approaches, including dietary interventions, behavior modification, and regular exercise, are common strategies to lower cholesterol levels. Current drug therapies (i.e., statins), despite the proven cholesterol lowering ability, with their high relative costs and associated side effects, are not viewed to be optimal long-term answers [1, 2]. It is more attractive to develop possible strategies and safer alternative therapies by modulating diet through probiotic interventions that could be promising and cost effective in lowering cholesterol [3].

Previous in vitro screening experiments revealed that strain PD2 (dosa batter isolate) and strain PH5 (handva batter isolate) were able to tolerate maximum bile concentration, produce bile salt hydrolase (BSH) enzyme, and could deconjugate bile salts like sodium taurocholate (ST) [4, 5]; they were identified as Lactobacillus fermentum by molecular typing methods. These potential probiotic strains were investigated for probiotic candidatures using obligatory tests as well as evaluated for safety aspect by performing amino acid decarboxylating activity (production of biogenic amines), hemolysis activity, antibiotic resistance, and gelatinase activity [4,5,6]. Both the strains showed negative response for all these tests indicating possible safe use for further investigations. Further, with respect to direct cholesterol assimilation (in vitro), PH5 showed a maximum reduction of 76.85% followed by PD2 69.66% compared to control by utilizing human plasma as a source of cholesterol [4, 5]. Thus, the study was undertaken to explore the therapeutic potential of probiotic dahi made from two potent probiotic Lact. fermentum strains, i.e., PD2 and PH5 in a hyperlipemic healthy adult Wistar rat model, with a particular focus as biotherapeutics for the management of high cholesterol in Indian population.

Materials and Methods

Lact. fermentum PD2 [gene accession no.KR612224] and PH5 [gene accession no.KR612226] were isolated previously from the batter of traditional Indian fermented nondairy products [4,5,6]. Further, both these strains having potential probiotic attributes were selected for current investigation on the basis of their better abilities to lower cholesterol in in vitro trials. For the preparation of probiotic (curd) diet, overnight grown test cultures, i.e., Lact. fermentum PD2 and PH5 were pelleted at 5000 rpm for 30 min at 4 °C and washed twice with saline. The cell pellet of each culture was inoculated into sterile reconstituted skim milk and incubated at 37 °C to achieve curdling. The curd samples from each culture PD2 and PH5 were stirred and diluted to attain the final concentration 107 and 109 CFU/g as two define doses, respectively. The concentrations of the bacterial strains were maintained constant and checked by plating on selective (MRS) agar media over the fermentation time during each trial.

Experimental Animals

Healthy adult Wistar albino rats (Either sex), 4–6 weeks old weighing 200–250 g, were used for the present study. The animals were housed under well-controlled conditions of temperature (22 ± 2 °C) and humidity (55 ± 5%) for 12:12 h light-dark cycle. Animals had free access to conventional laboratory diet (purchased from Pranav Agro Pvt. Ltd.) and tap water ad libitum throughout the adaptation period. The protocol of the experiment in this thesis was approved by Institutional Animal Ethical Committee (IAEC) of Anand Pharmacy College (Protocol No: APC/2015-IAEC/1504).

Experimental Design and Diet

At the end of the adaptation period, animals were randomly selected, weighed, and divided into seven different groups with six animals in each group with their assigned diet as shown in Table 1. Adult Wistar albino rats of Group B (model), Group C (Standard) and other experimental groups (Groups D, E, F and G) were made hyperlipidemic by the oral administration of high-fat diet (Atherogenic diet) by mixing with regular pellet diet. The base composition of the atherogenic (hyperlipidemic) diet included cholesterol, cholic acid, coconut oil, sucrose, and normal laboratory diet 2%, 1%, 10%, 40%, and 47%, respectively.

Table 1 Experimental groups with their respective diet
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Rats of the experimental groups (Groups D, E, F, and G) were also fed with 2 ml probiotic (stirred curd) in two doses once daily in the morning through oral feeding/gastric incubation for 28 consecutive days. Control and Model groups received an equivalent amount of normal saline. Rats of Group C were treated with markedly available standard drug atorvastatin (10 mg/kg of their body weight) only to compare effect of treatment with our probiotic solutions. During the entire course of the experiment, the rats had free access to water and to the group specific diet (20 g/100 g body weight per day). The experiment was carried out for 4 weeks. Food intake of the animal was observed daily whereas the body weight was determined weekly. After the feeding period, the rats were euthanized and the weight of the visceral organs (Liver, Kidney, Spleen, Lungs and Heart) and Fat pad (mesenteric, perirenal, and epididymal white adipose tissues) was measured.

Assay for Serum Lipid

Blood samples of the animals of each group were collected at 0 day and the end of the experiment on 28th day. The overnight-fasted animals were given light anesthesia and blood was collected from the retro orbital plexus with capillary tubes. Each sample of serum was analyzed for serum total cholesterol (TC), serum high-density lipoprotein cholesterol (HDL-C), serum low-density lipoprotein cholesterol (LDL-C) and triglycerides (TG) using commercially available kits (Coral Clinical Systems, Goa) based on the enzymatic oxidation of these molecules. Atherogenic indexes (TC/HDL-C and LDL-C/HDL-C ratios) were then calculated [1, 2].

Assay for Liver Cholesterol and Triglycerides

After euthanasia, the livers from all groups were collected, perfused, rinsed with physiological saline solution, weighed, and homogenized. Liver tissues were extracted following standard procedure of Shakuto et al. [7] with slight modifications. An aliquot extract was supplied for lipids determinations by observing absorbance in UV–Visible spectrophotometer (UV-1601 Shimadzu, Japan) using standardized commercial kit (Coral Clinical Systems, Goa).

Histopathological Analysis

Rat livers from each group were dissected out and frozen rapidly to about −20 °C till use. All the tissues are then placed in neutral formaldehyde (10%), embedded in paraffin and processed into 5 μm sections for light microscopy according to the routine procedures. The sections were stained with hematoxylin-eosin (HE) for histological assessment [8].

Analysis of Fecal Samples for Fecal Sterol, Fecal Water, and Fecal Microflora

Fecal droppings were collected during the last 2 days of life of rats, and fecal neutral and acidic sterols were extracted with slight modifications as suggested by Xie et al. [2]. In the middle of the 4th weeks, rat feces were amassed, weighed, and dried at 80 °C in a vacuum drying oven until a constant weight was achieved within 24 hours, and then reweighed. Fecal water content was calculated as suggested by Lee et al. [9]. Fecal water content (%) = [(weight before drying-weight after drying)/weight before drying] × 100.

Each sample was homogenized using a mortar and pestle using sterile PBS. Subsequent tenfold serial dilutions of each sample were plated in duplicate as per the method of Xie et al. [2]. Eosin methylene blue (EMB) agar was used for Escherichia coli, Violet red bile agar was utilized for coliforms, and MRS agar was used for total lactobacilli. Plates of total lactobacilli were incubated anaerobically at 37 °C for 48 h, while plates for the enumeration of E. coli and coliforms were incubated aerobically at 37 °C for 48 h. The numbers of colony-forming units (CFU) are expressed as log10 CFU per g.

Results and Discussion

The in vivo effect of two nondairy origin potent probiotics Lact. fermentum (PD2 and PH5) was analyzed to determine how their cholesterol metabolism would affect hyperlipidemic Wistar albino rats as the model system in current approach.

Effect of Potential Probiotic on Anthropogenic Parameters

All the experimental rats utilized in this study appeared healthy throughout the whole feeding period of 28 days. Their body weight, fat pad, and food consumption were calculated and recorded for all the groups during the experimental period as indicated in Table 2.

Table 2 Anthropogenic parameters of rats during 28 day
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The initial body weight of rats showed no significant difference among groups. After 28 days of experimental period, the rats in group B (Model) exhibited an increasing trend in body weight (289.78 ± 13.9 g), probably as a consequence of their greater calorie intake, due to the greater energetic density of the high cholesterol diet compared with group A (Normal) which fed a regular diet. While feeding probiotic 107 and 109 CFU/ml of Lact. fermentum PD2 and PH5, other experimental groups (groups labeled D, E, F, G) showed an increase in the final body weight compared to their initial weight. Vijayendra and Gupta [10] reported Lact. acidophilus fermented dahi helped to improve body weight gain in rats. Food consumption was found significantly less (80.39 ± 4.16) in group B (Model) compared to group A (normal) (100.57 ± 10.29). All probiotic fed groups (D, E, F and G) exhibited equal food consumption efficiency ratio as shown in Table 3. Food efficiency % was calculated on the basis of weight gain (g) and mean food consumption.

Table 3 Effect of probiotic PD2 and PH5 on weight of visceral organs of rats
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After the rats were killed, fat pad (mesenteric, perirenal, and epididymal white adipose tissues) was separated and weighed. The weight of fat pad was observed to be significantly raised in group B (Model) 3.42 vs. 2.97 g. Group C (Standard) and group G (PH Test dose 2) demonstrated similar weight of fat pad (p > 0.05), 2.84 and 2.92 g respectively. Lowest fat pad weight was found to be observed in case of PD Test dose 1 (2.06 g).

Effect of Potential Probiotic on Weights of Various Organs

After the rats were killed, the visceral organs (liver, kidney, spleen, heart, lungs) were collected and weighed as shown in Table 3. The rats had the lowest liver weight in group A (Normal) while significantly highest liver weight in group B (Model). Although administrating probiotic dahi alters the liver weight. Group D (Lact. fermentum PD2 dose 1) significantly lowered liver weight as compared with the group B (Model). Group E, F, and G also lowered the liver weight, but without statistical significance.

There were no significant difference in the weight of hearts, lungs, and kidneys among the seven groups. Significant differences between spleen organs weight of group A (Normal) and group B (Model) control was observed, with insignificant changes in treatment groups.

Effect of Potential Probiotic Feeding on Serum Lipid Profile

Reduction of serum total cholesterol (TC) and LDL-C and increase in HDL-C may be fundamental treatment opinion for CVDS. The effect of dietary treatments (probiotic curd) on serum lipid profile (serum total cholesterol, triglycerides, HDL-cholesterol, LDL-cholesterol) has been recorded in Table 4.

Table 4 Serum lipid profile (mg/dl) of rats at the end of feeding period
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Serum Total Cholesterol (TC)

Blood serum total cholesterol (TC) levels in the seven groups are shown in Fig. 1. Before the experiment, TC showed no significant difference among groups. High-cholesterol diet could significantly increase serum TC levels (462.74 ± 5.62 vs. 72.17 ± 3.2 mg/dl) in group B (Model) compared to group A (Normal). Group C (Standard) administrated with atorvastatin averted the rise in cholesterol levels by bringing 22.95% reduction as compare to group B (Model).

Fig 1
figure 1

Effect of probiotic on (i) Serum total cholesterol (TC) level; (ii) Serum triglyceride (TG) level; (iii) HDL-C level; and (iv) LDL-C level. *Values with different superscripts differ significantly (p < 0.05)

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Probiotic test solution (Groups D, E, F and G) showed varying degrees of cholesterol lowering abilities in vivo. At the end of 4 weeks, compared with the group B (Model), both the strains showed significant (p < 0.05) reduction in serum TC levels as 58.58, 57.74, 55.6, and 67.21% in Groups D, E, F, and G, respectively. Thus, groups D, E, and F (PD Test dose 1, 2 and PH dose 1) showed comparable effect on cholesterol reduction, while group G exhibited maximum total cholesterol lowering potential (151.77 mg/dl vs. 462.74 mg/dl of model). In a study of Mohania et al. [11] concerning lipid profile by feeding LaVK2 Dahi on hypercholesterolemic rats, plasma total cholesterol level was decreased by 22.6% when compared to rats fed with buffalo milk.

Serum Triglycerides (TG)

High-fat diet administered group B (Model) showed significant (p < 0.05) rise in serum triglyceride levels, 122.35% compared to group A (Normal). Treatment with standard and potential probiotics significantly reduced the triglyceride levels, 66.3%, 65.26%, 50.5%, 70.69%, and 66.21%, respectively, in groups C, D, E, F, and G; compared to group B (Model) as depicted in Fig. 1. There were no significant differences (p > 0.05) among groups C (Standard), D (PD test dose 1) F (PH test dose 1), and G (PH test dose 2) as they showed nearly similar effects on serum triglyceride level of rats. Thus, based on the results we can assume, probiotics might be able to lower serum triglyceride level akin to standard cholesterol lowering drugs available in the market (i.e., atorvastatin).

Batish et al. [12] reported 23.26% and 21.09% reduction in serum TC and TG, respectively, while evaluating potential of probiotic strains in SD (Sprague-Dawley) rats. Further, Singhora et al. [13] reported 20.69% reduction in serum TG level by feeding fermented milk with Lact. gasseri Lg70.

High-Density Lipoproteins (HDL-C)

High-density lipoprotein accumulates surplus cholesterol that cholesterol metabolizing cells cannot utilize. Thus, it’s an important factor to control arteriosclerosis. Atorvastatin administered in the group C (Standard) raised the HDL level in serum by 51.95% (highest compared to each group) compared to group B (Model). Moreover, group B (Model) showed 4.49% reduction (p > 0.05) when compared with group A (Normal). Furthermore, three groups, D, F, and G, exhibited (p > 0.05) no effect compare to group B (Model). Whereas, group E fed with LACT. fermentum PD2 showed significant decrease compared to Model and other probiotics (Fig. 1). Thus, treatment with probiotic did not show obvious influences on HDL levels in this study.

Similarly, Tamai et al. [14], Ibrahim et al. [15] and St-Onge et al. [16] did not observe any variation in the level of blood HDL cholesterol in rats or human by feeding fermented milk, milk yogurt, and soy yogurt. Lact. plantarum MA2 isolated from Tibetan kefir grains did not influence serum HDL level in rats [17].

Low Density Lipoproteins (LDL-C)

LDL-C is the main component of serum cholesterol inducing hypercholesterolemia. The elevated serum LDL cholesterol levels induced by feeding high-cholesterol diets were reduced in the PD2 and PH5 strains treated groups. Atorvastatin showed significant reduction (p < 0.05) 33.24% (184.76 ± 9.91 vs. 276.74 ± 7.42) in LDL level as compared to group B (Model). Potential probiotics (treatment groups D, E, F, and G) demonstrated significant (p < 0.05) decline in the LDL levels, 51.07%, 49.73%, 57.96%, and 63.25%, respectively, as compared to the group B (Model). As shown in Fig. 1, we can postulate that both PD Test dose 1 and 2 treated groups (D and E) exhibited comparable effect in LDL reduction 135.42 and 139.14 mg/dl, respectively. PH Test dose 1 (Group F) significantly diminished LDL (116.36 mg/dl vs. 276.74 of Model). Among all probiotic fed groups, PH Test dose 1 (Group G) was able to reduce maximum LDL (101.72 mg/dl) compared to group B (Model) 176.74 mg/dl.

Similarly, supplementation of diet with Lact. plantarum LP91 and LP21 resulted in significant reduction in LDL-C values by 38.13 and 21.42%, respectively [12]. In a recent study by Singhora et al. [13], test group which was fed on milk fermented with Lact. gasseri Lg70, a 49.54% significant reduction in LDL-C was observed compare to positive control (HD diet) group.

Atherogenic Index

TC/HDL-C and LDL-C/HDL-C ratios are recognized as two important signs of CVDs risk and were reduced in rats supplemented with LAB [1, 18]. The higher ratios are, the greater the CVD risk is. Therefore, reduction in serum TC, LDL-C, and increase in HDL-C may be an imperative treatment alternative for CVDs.

In the present study, as shown in Fig. 2, AI of the group B (Model) showed sharp increase after 4 weeks of feeding on the hyperlipidemic diet in comparison with that of group A (Normal control). All the four groups fed with probiotic dahi (D, E, F and G) including group C (Standard) exhibited significantly lower TC/HDL-C and LDL-C/HDL-C ratios when compared with hypercholesterolemic control group (Model) proposed their potentiality in cholesterol metabolism. Among all probiotic fed groups, Lact. fermentum PH5 fed groups (F and G) exhibited maximum potential to bring down atherogenic index by affecting TC/HDL-C and LDL-C/HDL-C ratios compared to PD2 fed groups (E and F). PH5 Test dose 2 has significantly high ability to reduce TC/HDL-C (3.58 vs. 11.18) and LDL-C/HDL-C (2.35 vs. 6.59). Batish et al. [12] also reported that AI for the probiotic (Lact. plantarum) treatment groups [HD91 (2.41) and HD21 (3.10)] decreased significantly when compared with the hypercholesterolemic control group (4.24).

Fig 2
figure 2

Effect of probiotic on atherogenic index. *Values with different superscripts differ significantly (p < 0.05)

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Effect of potential Probiotic feeding on Liver lipid profile

Liver total cholesterol (Liver TC)

Liver TC (Table 5) levels differed significantly among all the seven groups. The liver TC levels of rats fed a high-cholesterol diet group B (Model) had significantly increased (p < 0.05) compared to 163.39% with group A (Normal). Standard drug atorvastatin prevented the rise in liver cholesterol levels with reduction of 33.88% (235.4 ± 7.72 vs. 356 ± 9.73) as oppose to the group B (Model).

Table 5 Liver lipid profile (mg/dl) of rats at the end of feeding period
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PD test dose 1 and 2 (Group D and E) administration resulted in decreasing the liver cholesterol level (p < 0.05) by 24.97 and 16.34%, respectively. PH Test dose 1 (Group F) showed comparable effect with PD Test dose 1 (Group D) as shown in Fig. 3. While, Group C (Standard) treated with atorvastatin and PH Test dose 2 (Group G) showed maximum besides comparable potential to diminish liver cholesterol in rats. Similarly, fermented dairy product isolate NS12 (Lact. delbrueckii sp. bulgaricus) was reported effective to reduce 34.58% liver cholesterol [8]. The effect of dietary treatments (probiotic curd) on liver lipid profile (serum total cholesterol, triglycerides) has been recorded in Table 5.

Fig. 3
figure 3

Effect of probiotic on (i) Liver-TC level, and (ii) Liver-TG level. *Values with different superscripts differ significantly (p < 0.05)

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Liver Triglycerides (Liver TG)

The group A (Normal) displayed the lowest liver TG level and Model group (B) displayed the highest liver TG level. Atorvastatin treatment resulted in significant decline of 50.58% in the liver triglycerides value compared to group B (Model). All the four experimental groups (HC + PD2) and (HC + PH5) had lower levels of liver TG than HC group (p < 0.05), and there was no obvious differences among the four groups. All four experimental groups D, E, F, and G, fed with probiotic dahi, showed decrease in liver triglyceride values 31.57, 34.31, 41.82, and 44.96%, respectively, as shown in Fig. 3. Group C (Standard), and probiotic Lact. fermentum PH5 Test dose 2-treated group G demonstrated comparable results in terms of liver triglyceride reduction. The lower level of hepatic cholesterol and triglyceride content could reduce the rate of conversion of LDL to LDL-C particles, and result in the decline in serum LDL-C concentration, that might be another reason for the effects of our two strains to lower LDL-C level.

Supplementation with either of the two strains was effective in reducing serum TC, LDL-C, and TG concentrations in rats compared to those fed the same high-cholesterol diet but without LAB supplementation. In particular, these effects were more evident with Lact. fermentum PH5 fed groups. Our results confirm those described in previous reports [1, 8, 11, 12, 19].

Histopathology Analysis

Fig. 4 illustrated the effects of Lact. fermentum strains PD2 and PH5 on hepatic steatosis. Hematoxylin-eosin (HE) staining, a semi-quantitative method showed the differences in liver tissue structures and lipid accumulation of all groups.

Fig 4
figure 4

Histopathology of liver A: group A (Normal diet); B: group B (Model group-high cholesterol diet); C: High-cholesterol diet + PD2 probiotic dahi; D: High-cholesterol diet + PH5 probiotic dahi

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The rat liver of group A (Normal) had a well-organized structure as there was no fatty vacuolization in liver cells. The liver tissue in the group B (Model) had a moderate degree of vacuolization and increased lipid deposition in the cytoplasm. Hepatocyte steatosis was obviously alleviated by feeding probiotic dahi containing PD2 and PH5 strains compare to group B (Model). The LAB-treated rats exhibited overall normal gross liver appearances but high fatty vacuolization in Model group. Similarly, liver lipid deposition was evaluated in a semi quantitative manner by Xie et al. [2].

Fecal Sterol Analysis

Many probiotic strains could augment fecal elimination of bile acids through bile acid deconjugation, and this may alter the cholesterol synthesis pathways and resulted in a decrease of serum cholesterol concentration [20]. We found more bile acids released in the feces as well as highest fecal cholesterol especially in those rats fed with PH5 groups (F and G).

PH5 Test dose 1 released highest fecal cholesterol (52.26 mg/day) among other probiotic fed groups (D, E, and F) as well as group C (Standard). Moreover, both PH5-treated groups (F and G) liberated highest bile acid in the feces of rats 30.05 and 30.43 mg/day, respectively followed by group E (23.39 mg/day) and group D (20.18 mg/day). The lower fecal cholesterol and bile acids in the feces of rats of group B (Model) might be due to deposition or absorption of lipids in tissues as well as blood (Fig. 5). Overall, Lact. fermentum PH5 seems to be more effective in fecal cholesterol reduction and liberation of free bile acids through effective bile deconjugation mechanism. Similar experiment was performed by Xie et al. [2] using Lact. fermentum M1-16 feeding in hyperlipidemic rats; and reported 30.2% (37.33 mg/day) fecal cholesterol with 19% (19.29 mg/day) fecal cholic acid. Similar observation was reported by Batish et al. [12] as fecal cholic acid excretion increased throughout the experimental period in the probiotic treatment groups HDCap91, HD91 and HD21.

Fig 5
figure 5

(i) Analysis of fecal water content, (ii) Effect of probiotic on fecal cholesterol and bile acid (4th week). *Values with different superscripts differ significantly (p < 0.05)

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Fecal Water Content Analysis

Since the fecal water content can be exploited as an index of fecal elimination, our observations recommend that these two LAB strains have laxative potential and may stimulate bowel movements. As a result, the transit time for cholesterol absorption in the intestine might also be reduced.

Fecal water content data collected at the forth week is shown in Fig. 5. However, at the end of the feeding period, highest fecal moisture content was found in Lact. fermentum PH5 fed groups (F and G) as 67.01 and 65.88, respectively (p < 0.05). The results were comparable (p > 0.05) with group C (Standard) who demonstrated 62.1% moisture content in the feces of rats. PD2 Test dose 2 also exhibited 59.72 percent moisture in feces of rats. In a similar study by Xie et al. [2], probiotic solutions having Lact. plantarum 9-41-A and Lact. fermentum M1-16 exhibited 63% and 62% fecal moisture content, respectively, compare to their Model (53%).

Fecal Microbiological Analysis

Intestinal microbiota plays an important role in modulation of host energy and lipid metabolism [21]. The effects of probiotic lactobacilli on different bacterial counts in feces had been investigated previously by different research groups [22, 23]. The overall mean fecal bacteria counts at 0 and 28 days obtained from different experimental groups are recorded in Table 6 revealed the effects of the different diets and LAB supplementation on the rat intestinal bacterial flora.

Table 6 Effects of probiotic feeding on fecal bacterial population
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The rats fed on Lactobacillus strains as a dietary adjunct maintained a high level of lactobacilli (probiotic Lact. fermentum PD2 and PH5) in the fecal samples throughout the experimental period compared with the control groups. Total lactobacilli ranged from 8.47 to 9.94 CFU/g of feces in all the experimental groups. We also found Coliforms were significantly increased in high-cholesterol diets fed rats, and PD2 and PH5 strains restored the changes. Total coliforms count ranged from 6.03 to 7.24 CFU/g among all the treatment groups. Thus, increase in the number of fecal lactobacilli indicates their effective colonization in rat intestinal tract besides decrease in coliforms. However, rats supplemented with the diet containing PH dose 1 and 2 (Groups F and G) showed a marginal decrease in fecal E. coli counts compared with their counts on 0 day. Comparable fecal bacterial counts were obtained by Batish et al. [12] while studying hypercholesterolemia in SD rats using BSH producing Lact. plantarum. Hu et al. [8] reported relatively similar data while studying lipid metabolism in Lactobacillus strains, i.e., NS5 and NS12.

Conclusion

Present study demonstrated that animals fed with probiotic dahi (i.e., fermented with potential probiotic strains PD2 and PH5) might be useful in reducing hypercholesterolemia induced by the consumption of the atherogenic diet. At the outset, the probiotic dahi reduced plasma cholesterol and triglycerides suggesting that the effects of the probiotic dahi might have commenced at the level of the GI tract, inhibiting absorption of these lipids. In addition to reducing hepatic cholesterol and hepatic triglycerides, aortic tissue lipids were also decreased by ingestion of the probiotic dahi. Furthermore, the indigenous strains have also shown to possess bile salt hydrolase (BSH) activity in vitro and thereby suggesting that this BSH activity could be responsible for lowering the plasma total cholesterol concentration. Our study had some limitations, we used Wistar rats, and the effects observed in animal model may not be exactly the same in human. However, well-designed placebo-controlled clinical trials need to be conducted to validate the efficacy and safety of the strain and its use in the management of high cholesterol in humans.