Effects of Administering Fungi Isolated from Poultry Feed on Some Physiological and Productive Parameters in Rose Chickens in Several Regions of the Karbala Governorate

Mohanad Fadhl Hussain Al-Musodi*, Zeinab A.M. Al-Tememe, Mustafa Hadi Hamid

Department of Animal Production, College of Agriculture, University of Kerbala, Iraq.

Received | February 09, 2025; Accepted | April 27, 2025; Published | May 26, 2025

*Correspondence | Mohanad Fadhl Hussain Al-Musodi, Department of Animal Production, College of Agriculture, University of Kerbala, Iraq; Email: [email protected]

Citation | Al-Musodi MFH, Al-Tememe ZAM, Hamid MH (2025). Effects of administering fungi isolated from poultry feed on some physiological and productive parameters in rose chickens in several regions of the Karbala Governorate. J. Anim. Health Prod. 13(2): 393-401.

DOI | https://dx.doi.org/10.17582/journal.jahp/2025/13.2.393.401

ISSN (Online) | 2308-2801

Copyright: 2025 by the authors. Licensee ResearchersLinks Ltd, England, UK.

This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

Introduction

The economic cost of feed significantly influences the growth of the livestock sector and constrains animal production. Therefore, rationing feed while maintaining production quality and quantity is crucial. Numerous researchers have explored the use of low-cost feed additives in minimal ratios to enhance the quality and quantity of meat, eggs, and other products by improving feed quality with additives not suitable for human consumption (Ayoola et al., 2024; Classen and Coppe, 1998). Agricultural animals, including broilers, can convert low-quality feed unsuitable for humans into high-quality animal protein, with feed accounting for about 75% of total poultry farming costs (Atteh, 2004).

Poultry can be exposed to various pathogens, including fungi and their toxins, through the consumption and digestion of contaminated feed especially yellow corn (Canas and Aranda, 1996; Al-Tememe et al., 2025). Fungal contamination of poultry feed is a global issue, leading to altered blood parameters, impaired liver function, reduced growth and development, and significant economic losses for poultry meat and egg producers (Che et al., 2011). Additionally, fungi can also be found in broiler drinking water and airborne dust (Ibrahim et al., 2021). Aspergillus species can affect birds of any age and in any environment (Abd El-Ghany, 2021).

The high concentration of fungi or their toxins in the feed leads to an increase in the percentage of halogens, a decrease in red blood cells, egg production, deterioration of the nutritional conversion rate, inhibition of immune system function, decreased response to vaccines, and pathological changes in the liver and other parts of the bird’s body (Kamalavenkatesh et al., 2005). Studies have pointed to a scientific fact that the higher the concentration of mycotoxins, the activity the immune system is inhibited, the effectiveness of phagocytic cells decreases, and lymphocytes decrease (Fung and Clark, 2004). Aspergillus flavus produces mycotoxins specifically aflatoxins B1, B2, G1, and G2 which commonly contaminate cereals such as wheat, yellow corn, and soybeans at various stages of production (Cole and Richard, 1993). Among these, aflatoxin B1 is considered the most harmful to poultry, leading to symptoms like reduced appetite, poor weight gain, decreased feed intake, impaired feed conversion ratio, and even mortality (Girish and Devegowda, 2006). The liver is the primary target organ affected by mycotoxins (Miazzo et al., 2000), and mycotoxin contamination has long been a challenge in the poultry industry (Kubena et al., 1993). In contrast, Aspergillus niger exhibits biostimulant properties and is commonly found in nature and easily cultured in laboratories (Piva et al., 1995). It is a valuable source of bioactive compounds such as citric, gluconic, and itaconic acids, and it produces beneficial enzymes like protease, cellulase, xylanase, and phytase, which support bird health and can enhance the economic efficiency of poultry farming (Nadumane et al., 2016).

As a bio-enhancer, proprebiotics (when alive) and prebiotics (when inactivated) support the growth of beneficial gut bacteria in poultry and help birds utilize the physiologically active secondary metabolites produced by fungi (Hong et al., 2004). Previous studies have shown that supplementation with Aspergillus niger improves digestive health and reduces populations of E. coli and Salmonella in birds (Saleh et al., 2017).

The aim of the study was to make a comparison between the effect of two types of fungi that were isolated from a poultry feed in Karbala on some productive and physiological parameters in broiler.

Materials and Methods

This study was conducted at the Animal Field, Faculty of Agriculture, Karbala University, using 180 broiler chickens of the Rose breed. The experimental birds were reared and handled in line with all ethical standards and study protocols were approved by the University of Kerbala. The birds were randomly assigned to three equal groups, with each group further divided into six replicates of 10 chicks each. All birds were maintained on a standard balanced diet. The second group received a 1 ppm solution of Aspergillus flavus, while the third group was dosed with a 1 ppm solution of Aspergillus niger. The first group, which received no fungal treatment, served as the control.

Isolation and diagnosis of fungi used in the study

A. flavus and A. niger fungi were isolated from poultry feed pellets in Karbala, Iraq, using a modified method described by Ali et al. (1991) and identified according to the procedure outlined by Kayode and Sani (2008).

Productive parameters

Body weight and feed efficiency were measured weekly, starting from the seventh day of the experiment. Each chick was individually weighed on the first day, and this procedure was repeated every weekend throughout the trial. Live body weight and weight gain were calculated using the method described by Al-Musodi et al. (2024), by subtracting the bird’s weight at the start of the week from its weight at the end of the week.

Feed consumption was determined based on the actual feed intake of each group, following the method of Tarrage and Puchal (1977). It was calculated by subtracting the amount of feed remaining at the end of the week from the amount provided at the beginning.

The feed conversion ratio (FCR), which indicates feed efficiency, was calculated using the following formula:

FCR = Feed consumed (g)/Average weight gain (g)

This value represents the grams of feed required to produce one gram of body weight gain (Al-Musodi et al., 2024).

Blood collection

At the end of the rearing period, blood samples were collected from the birds’ jugular veins. A total of 5 mL of blood was drawn from each bird and divided into two tubes. One tube contained an anticoagulant for the analysis of white blood cells, hemoglobin, packed cell volume (PCV), and leukocyte counts. The second tube, left without anticoagulant, was used to collect serum for the measurement of lipid profile, liver enzyme activity, and blood protein levels.

Hematological analysis

Bird blood cells were diluted using Natt and Herrick solution and analyzed using the hemocytometer counting method, following the procedure described by Campbell (1988). For red blood cell (RBC) counting, 0.5 mL of blood was drawn and diluted with the solution to the 101 mark, resulting in a 1:200 dilution. For white blood cell (WBC) counting, the same volume of blood was diluted to the 11 mark, yielding a 1:20 dilution. The cells were then counted using standard hemocytometer procedures.

Differential leucocyte counts were performed by preparing a blood smear. A drop of blood was placed on one end of a clean glass slide, and another slide held at a 45° angle was used to spread the blood into a thin film. After air-drying, the smear was stained with Wright’s stain, gently rinsed, and dried again. The slide was examined under an oil immersion lens, and 100 leukocytes were counted. The different types of white blood cells were identified based on morphological characteristics, as per the method of Burton and Harrison (1969).

Packed Cell Volume (PCV) was measured by collecting blood in heparinized capillary tubes after puncturing the wing vein. One end of the tube was sealed with artificial clay, and the tube was centrifuged at 12,000 rpm for 5 minutes using a hematocrit centrifuge. PCV was then determined using a hematocrit reader, following Campbell’s (1988) method.

Hemoglobin concentration was estimated using Drabkin’s solution, which converts hemoglobin to cyanmethemoglobin. A volume of 20 µL of blood was mixed with 5 mL of Drabkin’s solution and allowed to react for 5 minutes. The mixture was then centrifuged at 2,500 rpm for 5 minutes, and absorbance was read at 540 nm using a spectrophotometer. The spectrophotometer was zeroed with Drabkin’s solution, in accordance with Varley and Bell (1980).

Determination of the total protein level in the blood

The Biuret Method, described by Wotton (1964), was applied to measure the total protein concentration in the blood. This method is based on the interaction between copper ions in the biuret reagent and protein peptides in an alkaline medium, forming a violet-colored complex. The Rodkey (1965) method was employed to measure the albumin concentration in the blood. To determine the globulin concentration, the total protein level was subtracted by the albumin concentration, as described by Ghanim et al. (2016).

Determination of lipid profile

Serum total cholesterol concentration (mg/dL) was measured using a commercial kit (Spinreact, Spain) following the procedure outlined by Allian et al. (1974). Triglyceride levels were also determined with a commercial kit (Spinreact, Spain), following the method described by Fossati and Prencipe (1982).

High-Density Lipoprotein (HDL-C) concentration (mg/dL) was measured using a commercial kit (Spinreact, Spain), as per the procedure outlined by Tietz (1995). Similarly, Low-Density Lipoprotein (LDL-C) concentration (mg/dL) was calculated using the method of Buritus and Ashwood (1999) with the following equation:

LDL-C concentration (mg/dL)= Total cholesterol - (HDL-C + vLDL-C)

Very Low-Density Lipoprotein (vLDL-C) concentration (mg/dL) was measured according to Buritus and Ashwood (1999), using the following formula based on triglyceride levels:

vLDL-C concentration (mg/dL) = Triglycerides/5

Liver function enzymes

Alanine Aminotransferase (ALT) activity (U/L) was measured using the method described by Wroblewski and Ladue (1956). Aspartate Aminotransferase (AST) activity was quantified according to the procedure outlined by Jain (1986).

Renal function

The concentration of urea in the blood of meat chicks was determined using the method described by Chawla (1999), while the blood creatinine level was measured according to the method of Naseem et al. (2018).

Statistical analysis

Statistical analyses in this study were performed using SPSS 15.0. ANOVA was used to assess the significance of the main effects and interactions in the data. The Duncan Multiple Range Test was employed to compare the means. The significance threshold was set at P < 0.05 (SAS, 2010).

Results and Discussion

Live body weight

It is noted from the Table 1 that the weights of the chicks in the third group were significantly higher (p<0.05) than those of the control group and the second group in the fourth week of life. Additionally, the weights of the control group were significantly higher (p<0.05) than those of the second group in the same week. However, the chicks in the first and third groups showed similar weights.

Body weight gain

The chicks of the third group were significantly superior (p<0.05) in terms of weekly weight gain over the chicks of the second group in the last week of the study (Table 2).

Feed intake

It is noted that there were no significant differences (p>0.05) in the amount of feed intake between the chicks of the study groups, as shown in Table 3.

Feed conversion ratio (g/ week)

The addition of A. niger to the broiler chicks resulted in a significant (p<0.05) improvement in the feed conversion rate compared to the control and second groups in the forth week of the study, as shown in Table 4.

Blood cell parameters

The third group significantly (p<0.05) outperformed the second group in the number of red and white blood cells, hemoglobin levels, and the percentage of packed cell volume (PCV), while the control group surpassed the second group in the number of white blood cells, as shown in Table 5.

Differential leucocyte count

The results of the differential white blood cell count showed a significant (p<0.05) higher percentage of heterophils in the control group compared to the second group. In contrast, the second group significantly outperformed the control group in the percentage of lymphocytes and eosinophils, as shown in Table 6.

 

Table 1: The effect of treatment on weekly body weight (g/week) of broilers.

Groups/ week	1st week	2nd week	3rd week	4th week	5th week
Control	185.57±5.91	518.07±36.21	882.77±34.61	1482.08±31.23b	2150.4±60.1a
Group 2 (A. flavus)	174.81±20.11	489.01±30.13	810.11±13.54	1314.12±30.17c	1892.15±58.07b
Group 3 (A. niger)	١٧٨.٨٤±8.73	532.24±31.02	982.14±13.15	1565.9±9.11a	2254.9±71.02a
LSD	45.04	126.11	94.12	82.91	201.4
P value	N.S	N.S	N.S	<0.05	<0.05

 

Table 2: The effect of treatment on weekly weight gain (g/week) of broilers.

Groups/ week	1st week	2nd week	3rd week	4th week
Control	332±50	364.70±25.40	599.30±37.41	668.40±24.64ab
Group 2 (A. flavus)	314.2±30.71	321.10±32.18	504±61.60	578.04±99.34b
Group 3 (A. niger)	353.40±20.14	449.90±40.01	583.74±17.90	689.05±29.11a
LSD	90.06	111.91	154.12	104. 1
P value	N.S	N.S	N.S	<0.05

 

Table 3: The effect of treatment on feed intake (g/Week).

Groups/ week	1st week	2nd week	3rd week	4th week
Control	509.70±40.10	657.40±23.41	679.80±24.10	936.50±37.39
Group 2 (A. flavus)	440.81±70.10	627.97±25.01	677.80±29.60	805.17±36.06
Group 3 (A. niger)	478.40±21.17	624.40±27.51	807.10±27.31	808.19±61.40
LSD	97.11	88.40	590.10	163.90
P value	N.S	N.S	N.S	N.S

 

Table 4: The effect of treatment on feed conversion ratio (g/week).

Groups/ week	1st week	2nd week	3rd week	4th week
Control	1.53±0.4	1.80±1.24	1.13±0.10	1.40±0.32a
Group 2 (A. flavus)	1.40±0.44	1.95±2.30	1.34 ±0.17	1.39±1.03ab
Group 3 (A. niger)	1.35±0.90	1.38±1.13	1.38±3.21	1.17±0.33b
LSD	0.713	0.631	0.591	0.291
P value	N.S	N.S	N.S	<0.05

 

Table 5: The effect of treatment on blood picture.

Groups/ week	Red Blood cells Cellx106	White blood cells Cellx103	Hemoglobin (Hb) g/dl	Packed cell volume (PCV) %
Control	45.26±0.7ab	50.82±1.02ab	8.80±0.12b	29.48±0.50a
Group 2 (A. flavus)	42.40±1.04b	49.70±0.75b	7.81 ±0.11c	25.40±1.14b
Group 3 (A. niger)	46.41±1.27a	54.75±0.43a	9.91±0.07a	31.10±0.17a
LSD	3.11	3.017	0.640	2.41
P value	<0.05	<0.05	<0.05	<0.05

 

Table 6: The effect of treatment on differential Leucocyte count (%).

Groups/week	Heterophil	Lymphocyte	Eosinophil	Basophil	Monocytes
Control	27.91±0.12a	64.61±0.14b	0.65±0.11b	0.71±0.01	6.64±0.17
Group 2 (A. flavus)	25.17±0.41b	66.94±2.91a	0.78 ±0.13a	0.75±0.03	6.84±0.44
Group 3 (A. niger)	26.99±0.71ab	65.01±2.81ab	0.86±0.14a	0.75±0.04	6.75±0.30
LSD	1.12	2.04	0.11	0.170	1.301
P value	<0.05	<0.05	<0.05	N.S	N.S

 

Table 7: The effect of treatment on Lipid profile (µg/dl).

Groups/ week	cholesterol	Triglycerides	HDL	LDL	vLDL
Control	136.51±1.17a	49.55±1.14	23.31±1.09ab	102.11±1.14	9.91±0.83
Group 2 (A. flavus)	133.41±1.90b	48.56±1.81	22.50 ±0.70b	100.31±1.21	9.71±0.72
Group 3 (A. niger)	133.90±1.71b	54.50±3.09	24.61±1.90a	99.20±0.017	10.90±2.14
LSD	2.10	7.35	2.11	3.24	5.31
P value	<0.05	N.S	<0.05	N.S	N.S

 

Table 8: The effect of treatment on biochemical parameters.

Groups/ week	Total protein g/dl	Albumin g/dl	Globulin g/dl	AST u/l	ALT u/l	Urea mg/dl	Creatinine Mg/dl
Control	3.33±0.42b	1.42±1.14a	1.91±0.31ab	120.36±14.12b	401.34±3.74b	3.18±11.21	0.41±13.22b
Group 2 (A. flavus)	3.01±012b	1.31±0.43b	1.70±0.36b	129.31±13.98a	411.60±6.14a	3.21±14.2	0.47±17.12a
Group 3 (A. niger)	3.63±0.1a	1.44±0.4a	2.19±0.3a	122.14±15.0ab	404.19±3.8ab	3.27±20.1	0.43±21.0ab
LSD	0.857	0.10	0.411	8.51	5.314	2.150	0.055
P value	<0.05	<0.05	<0.05	<0.05	<0.05	N.S	<0.05

 

Lipid profile

Fungal treatments influenced the blood lipid profile, with the total cholesterol concentration significantly higher (p<0.05) in the control group compared to the second group. Additionally, the concentration of high-density lipoproteins (HDL) was higher in the third group compared to the second group, as shown in Table 7.

Biochemical parameters

As shown in Table 8, the concentrations of total blood proteins, albumin, and globulin in the third group were significantly higher (p<0.05) compared to the second group. Additionally, the concentrations of the enzymes AST and ALT were significantly higher (p<0.05) in the second group compared to the third group. The concentration of creatinine was significantly higher (p<0.05) in the second group compared to the control group. However, the concentration of blood urea was not significantly affected by the treatment.

Discussion

The improvement in the productive performance of broilers in the third group, as shown in Tables 1, 2, and 4, reflects the positive effects of A. niger on the gastrointestinal tract. This fungus enhances nutrient utilization particularly carbohydrates, polysaccharides, and enzymes by improving their digestion and releasing bioactive compounds that promote chick growth (Muhammed and Oloyede, 2010). A. niger also improves the digestion of complex substances due to its probiotic-like action (Muhammed and Oloyede, 2006), mediated by enzymes it secretes, such as pectinase, carbohydrase, lactase, invertase, and acid protease (Nagashima et al., 1999). These enzymes boost protein digestion and mineral absorption, both crucial for growth, weight gain, and overall health (Nelson and Cox, 2005). These findings align with those of Muhammed and Oloyede (2010), Omid et al. (2022), and Saleh et al. (2011), who reported increased broiler weights after A. niger supplementation.

The improved feed conversion ratio observed in the third group may also result from enhanced feed utilization due to A. niger’s enzymatic activity, consistent with previous findings in various animals (Muhammad et al., 2000; Omid et al., 2022; Kayode et al., 2008).

In contrast, the second group showed a significant (p<0.05) decline in productive traits compared to the control and third groups, likely due to the harmful effects of aflatoxins produced by A. flavus (Freire et al., 1996). This result agrees with earlier studies by Jeff-Agboola (2014), Nazarizadeh et al. (2019), and Khaleghipour et al. (2019), which reported negative impacts of aflatoxins on poultry performance.

Significant heterogeneity (p<0.05) was observed in the blood parameters, particularly a decrease in the second group, likely due to the effects of aflatoxins secreted by A. flavus (Anwar et al., 2001). Blood parameters are key indicators of an animal’s health, and reductions in hemoglobin, packed cell volume (PCV), and erythrocyte count have similarly been reported in layer hens fed A. flavus-contaminated feed (Fawaz et al., 2022). Aflatoxins are known to trigger inflammation and cellular damage, contributing to these hematological disruptions, as also observed in turkey chicks (Javed et al., 2005), with findings supported by Basmacioglu et al. (2005).

In contrast, Table 5 shows a significant increase (p<0.05) in blood parameters in the third group. These findings align with Al-Kassie et al. (2008), who reported that probiotics significantly enhanced hemoglobin levels in broilers, improving oxygen delivery and reducing hypoxia risk. A. niger may contribute to better oxygen saturation and support the development and protection of red blood cells and their hemoglobin content (Pavlidis et al., 2007). Similar results were reported by Hao et al. (2020), who observed a significant increase in blood parameters in chicks supplemented with A. niger at different dietary levels (0.25%, 0.5%, 0.75%, and 1%).

As for differential leukocyte counts, Table 6 indicates a significant increase (p<0.05) in lymphocyte percentage in the second group, which may reflect the immunological stress caused by aflatoxin exposure. Aflatoxins contribute to anemia, cellular damage, and reduced red and white blood cell counts, consistent with the findings of Jeff-Agboola (2014). The elevated eosinophil levels in the second group could be attributed to hypersensitivity or inflammatory responses triggered by aflatoxins or fungal elements (Betina, 1989; Oguz et al., 2000), further supported by Anjorin and Cyriacus (2014).

The concentration of high-density lipoproteins (HDL) significantly increased (p<0.05) in the third group compared to both the control and second groups (Table 6), suggesting the beneficial role of A. niger in lowering harmful cholesterol levels (Al-Kassie et al., 2008; Kim et al., 2003). This effect may be attributed to the fungus’s influence on the enzyme HMG-CoA reductase, which is critical in cholesterol biosynthesis and is known to be inhibited by fungal metabolites (Hajjaj et al., 2005). Similarly, Srinivasan et al. (2022) demonstrated reduced cholesterol levels in broilers supplemented with A. niger and probiotics, consistent with the findings of Saleh et al. (2011).

Further, A. niger may reduce hepatic cholesterol synthesis by suppressing liver enzyme activity and decreasing bile secretion (Tufan and Bolacali, 2017). Its role in enhancing short-chain fatty acid production in the intestine may also contribute to lower cholesterol formation in the liver (Tang et al., 2017). Increased HDL levels and reduced cholesterol could also stem from reduced lipid absorption in the intestine due to microbial activity stimulated by A. niger (Abul-Lateef et al., 2019; Awais et al., 2019). This may involve beta-fructan secretion by probiotic-associated fungi (Saleh et al., 2011; Hao et al., 2020).

In contrast, the decrease in cholesterol observed in the second group may result from the toxic effects of aflatoxins produced by A. flavus (Solis et al., 2019). Liu et al. (2018) similarly reported reduced cholesterol levels in broilers exposed to aflatoxins, likely due to impaired liver function and lipid metabolism.

The concentration of total protein, albumin, and globulin in the blood of the third group increased significantly (p<0.05) compared to both the second and control groups, as shown in Table 7. This improvement may be attributed to the role of A. niger in enhancing the utilization of organic compounds in the feed, particularly proteins (Abul-Lateef et al., 2019). In contrast, protein levels in the second group (A. flavus) were significantly lower (p<0.05) than those in the control group, likely due to the toxic effects of aflatoxins (Zou et al., 2023).

However, despite this reduction, the chicks’ vital functions were not severely affected, possibly due to the low fungal dosage and relatively short rearing period (35 days), which the liver may have managed without major dysfunction (Lafi et al., 2010). These findings are consistent with those of Yunus et al. (2011), who also observed minimal physiological disruption at lower aflatoxin exposure levels over short durations.

Fungi can influence liver cells in multiple ways, either negatively or positively, depending on the fungal species and the bioactive compounds or toxins they produce (Ozer et al., 2008). The significant increase (p<0.05) in liver enzyme levels observed in the second group (A. flavus) may be attributed to liver cell damage and increased cellular breakdown (Yunus et al., 2011). Exposure of broiler chicks to mycotoxins is known to reduce blood protein levels while elevating liver enzymes (Soares et al., 1989). In this study, the observed signs of anemia further suggest possible liver fibrosis, likely caused by aflatoxin exposure (Zou et al., 2023). These findings are consistent with Liu et al. (2018b), who reported decreased total protein and albumin levels, alongside increased liver enzyme activity, in broilers administered aflatoxins derived from A. flavus.

The elevated creatinine levels observed in the second group (A. flavus) may be attributed to the effects of aflatoxin secretion, which can cause damage to kidney tissues. This damage likely impairs the kidneys’ ability to effectively eliminate metabolic waste from the body (Khaleghipour et al., 2019). These findings are consistent with those of Ukoha et al. (2023), who also reported increased blood creatinine concentrations in birds fed aflatoxin-contaminated feed.

Conclusions

According to the study, A. flavus negatively affects broiler growth and health parameters, while A. niger shows beneficial effects. Therefore, A. niger may be further explored using molecular techniques before being applied in commercial poultry production.

Acknowledgment

I extend my gratitude to the Faculty of Agriculture, Karbala University and to the Department of animal production to facilitate the task of conducting this study and provide the necessary laboratories and facilities for it. I would also like to thank the Journal of animal health and production for reviewing this study and correcting errors in it.

Novelty Statement

This article is one of the most recent articles in the country where it was conducted.

Author’s Contribution

The researchers contributed to the completion of this work with regular group work.

Conflict of interest

The authors have declared no conflict of interest.

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