Effects of three feed disinfectants on microbial population and nutritional value of broiler diets

Abstract

Feed hygienic quality can influence growth performance, health status, and production efficiency in broiler chickens. This study evaluated three chemical feed disinfectants differing in active ingredients—formaldehyde (formalin), a formaldehyde–propionic acid blend (Termin Gold®), and a hydrogen peroxide–silver nanoparticle formulation (Nanosil Feed & Air®)—and compared direct addition versus premix application on feed microbial quality, nutrient composition, and broiler performance. A total of 480 one-day-old male Ross 308 broiler chicks were used in a 35-d trial and allocated to 6 dietary treatments (4 pens/treatment; 20 birds/pen): control (no disinfectant), Termin Gold® added to the final diet, Nanosil Feed & Air® added to the final diet, formaldehyde added to the final diet, formaldehyde applied as a premix (prepared 7 d before use), and Termin Gold® applied as a premix (prepared 7 d before use). Growth performance was recorded weekly. Chemical composition of starter (d 0–10), grower (d 11–24), and finisher (d 25–35) diets was analyzed. Microbiological evaluations included total bacterial count, fungal count, and coliform population. From week 2 onward, premix-applied formaldehyde and Termin Gold® improved BW gain, feed intake, and feed conversion ratio, particularly during the grower and finisher phases. Disinfectant treatment did not adversely affect proximate diet composition. All disinfectants reduced bacterial contamination in starter and grower feeds, with formaldehyde showing the greatest effect in grower diets. Fungal counts were largely unaffected, whereas formaldehyde-based treatments produced the lowest coliform counts during the grower and finisher phases. In conclusion, both active ingredient and application method influenced disinfection efficacy. Premix application of formaldehyde- and formaldehyde–propionic acid–based disinfectants improved feed hygienic quality and broiler performance without compromising nutritional value.

Introduction

Feed hygiene is a critical determinant of health, performance, and productivity in modern broiler production systems (Abdel Moneim et al., 2024). Feed serves as a primary interface between the production environment and the bird, and its microbiological quality can influence growth rate, feed efficiency, immune function, and mortality (Stanley and Bajagai, 2022). Microbial contamination of feed can cause nutrient degradation, reduced digestibility, gastrointestinal disorders, and infectious disease risk, leading to economic losses (Graziano et al., 2013). Contaminated feed may also transmit pathogenic microorganisms along the food chain and raise food safety concerns (Zentek et al., 2013). Therefore, controlling microbial contamination during feed manufacturing, storage, and on-farm use is an essential component of poultry management (Mgbeahuruike et al., 2025).

Thermal processing (e.g., pelleting) can reduce microbial loads through combined effects of heat, moisture, and pressure (Rúgeles et al., 2010). However, pelleting is not universally applicable because of infrastructure limitations or reliance on mash diets in some production systems (Stanga, 2010). Under these conditions, chemical disinfection of feed is a practical alternative (Mehmet Akif et al., 2006) that can suppress microbial growth and improve hygienic quality without heat treatment (Fraise et al., 2013).

Despite antimicrobial effects, performance responses to disinfectants are inconsistent (Garcia et al., 2007). Some studies report improved body weight gain and feed efficiency (Ricke et al., 2019), whereas others suggest that inappropriate product selection or exposure may reduce palatability or alter gut microbial ecology (Avila et al., 2023; Barikgugjlu et al., 2016). Disinfectants may also interact with nutrients and potentially affect their availability (Leeson and Summers, 2009). Application strategy can influence distribution uniformity and microbial inactivation (Avila et al., 2023; Sbardella et al., 2015; Dibner and Buttin, 2002).

Accordingly, the present study evaluated three feed disinfectants—formalin (formaldehyde solution), Termin Gold® (a formaldehyde–propionic acid blend), and Nanosil Feed & Air® (a hydrogen peroxide–silver nanoparticle formulation)—and compared direct addition versus premix application for their effects on feed microbial quality, proximate chemical composition, and broiler performance. We hypothesized that dietary chemical disinfection using formaldehyde, organic acid–formaldehyde blends, or silver-stabilized hydrogen peroxide would significantly decrease microbial contamination in mash diets without deteriorating dietary proximate nutrient composition or broiler performance.

Materials and methods

Study design and ethical approval. The experiment was conducted during the winter of 2024 at the poultry housing facility of the Aminabad Institute of Research, Faculty of Veterinary Medicine, University of Tehran. A completely randomized design (CRD) was employed, consisting of 6 dietary treatments with 4 replicate floor pens per treatment (n = 24 pens). Sample size and replication were determined to achieve sufficient statistical power (α = 0.05, power = 0.80) to detect meaningful differences in growth performance and microbiological parameters. All procedures involving animals were approved by the Institutional Animal Care and Use Committee (IACUC) of the University of Tehran (IR.UT.VETMED.REC.1404.021) and complied with relevant ethical guidelines.

Birds, housing, and management conditions. A total of 480 one-day-old male Ross 308 broiler chicks were obtained from a commercial hatchery, individually weighed, and randomly allocated to 24 floor pens (120 × 120 cm, providing a standard stocking density with 20 birds/pen). Pens were bedded with double-layered corrugated cardboard paper over the concrete floor. The windowless house was equipped with a cross-ventilation system ensuring uniform airflow. House temperature and relative humidity (RH) were continuously monitored; the Temperature-Humidity Index (THI) was maintained within the optimal thermal comfort zone for broilers (initial brooding temperature of 32 to 33°C, gradually decreased to 21 to 22°C by wk 5; RH maintained between 45% and 65%). During the first 72 h, continuous lighting (24L:0D) was provided, followed by a lighting schedule of 23 h of light and 1 h of darkness (23L:1D, from 23:00 to 24:00) until the end of the trial. Birds had ad libitum access to feed and fresh water throughout the 35-d period.

Medication and vaccination program. No prophylactic antibiotics, growth promoters, or coccidiostats were included in the experimental diets or drinking water. The birds were vaccinated using the routine vaccination schedule in the region as follows: Cevac® Vitabron L and Cevac® Ibird were administered in combination by coarse spray on day 1; Izovac H120 CLONE was administered by eye drop and NewFluRazi® oil-based inactivated vaccine by subcutaneous injection on day 8; Avishield® ND Lasota was administered via drinking water on day 13; GUMBOKAL® IM SPF was administered via drinking water on day 18; Hipraviar® CLON was administered via drinking water on day 20; and Avinew® was administered by coarse spray on day 27.

Diets and experimental treatments. Experimental diets were formulated in three phases: starter (d 0–10), grower (d 11–24), and finisher (d 25–35; Table 1). Diets were formulated to meet nutrient requirements for Ross 308 broilers. The six treatments were: (T1) control (no disinfectant); (T2) Termin Gold® (Rooyan Darou Pharmaceutical Co., Semnan, Iran) was added to the final diet at 2 kg/ton directly in the main mixer; (T3) Nanosil Feed & Air® (Kimiafaam Pharmaceutical Company, Tehran, Iran) was applied as a 2% solution at 20 L/ton via direct mixing to the final diet in the main mixer; (T4) formalin (formaldehyde solution)(Pars Petrochemicals, Asalouyeh, Iran) was added to the final diet at 2 kg/ton directly in the main mixer; (T5) formalin was applied in a premix preparation at 2 kg/ton of final feed diet(prepared 7 d before incorporation in the final diet); and (T6) Termin Gold® was applied as a premix at 2 kg/ton of final feed formulation (prepared 7 d before incorporation in the final diet). For the latter treatments (T5 and T6), the disinfectants were mixed with the premix formula in a small micromixer. The premix formula was stored for a week and thereafter was added to the main mixer for the preparation of final formulation. Because of the liquid formulation of Nanosil Feed & Air®, its application in the premix preparation was excluded from the study. All products were used according to the manufacturer instructions, and mixing procedures were standardized.

Table 1.

Ingredient composition and calculated nutrient content of experimental diets.

Ingredient	Starter (0–10 d)	Grower (11–24 d)	Finisher (25 d–slaughter)
Corn	49.38	53.69	59.67
Soybean meal	43.32	39.19	33.64
Vegetable oil	3.310	3.620	3.420
Calcium carbonate	1.190	0.700	0.470
Salt	0.300	0.300	0.300
Chin-e Chin concentrate⁎	2.500	2.500	2.500
Calculated nutrient composition
Nutrient	Starter	Grower	Finisher
Metabolizable energy (kcal/kg)	2975	3050	3100
Crude protein (%)	23.00	21.50	19.50
Calcium (%)	0.95	0.75	0.65
Available phosphorus (%)	0.50	0.42	0.36
Methionine + cysteine (%)	1.00	0.92	0.86
Lysine (%)	1.33	1.22	1.10
Sodium (%)	0.18	0.18	0.18
Chloride (%)	0.21	0.21	0.21
Arginine (%)	1.28	1.26	1.28
Tyrosine (%)	0.89	0.89	0.72
Tryptophan (%)	0.23	0.23	0.23
Linoleic acid (%)	1.84	1.18	1.18

^⁎The Chin-e Chin concentrate contained (per kg): metabolizable energy, 1500 kcal; crude protein, 15.30%; methionine + cysteine, 7.28%; lysine, 3.50%; threonine, 1.00%; available phosphorus, 11.30%; calcium, 23.35%; sodium, 2.14%; chloride, 4.25%; zinc, 44000 mg; manganese, 48000 mg; copper, 6400 mg; iron, 8000 mg; iodine, 500 mg; selenium, 120 mg; choline, 80000 mg; vitamin A, 4000000 IU/kg; vitamin D3, 1800000 IU/kg; vitamin E, 26000 IU/kg; vitamin K3, 1000 mg/kg; vitamin B1, 1000 mg/kg; vitamin B2, 2600 mg/kg; vitamin B3, 24000 mg/kg; vitamin B5, 7200 mg/kg; vitamin B6, 1280 mg/kg; vitamin B9, 760 mg/kg; vitamin B12, 8 mg/kg; biotin, 72 mg/kg; antioxidant, 32000 mg/kg.

Chemical analyses. Proximate composition of the diets was determined according to official AOAC methods, including dry matter (AOAC 930.15), crude protein (AOAC 990.03), ether extract (AOAC 920.39), crude fiber (AOAC 978.10), and ash (AOAC 942.05).

Microbiological analyses. For each treatment and feeding phase, feed samples were collected aseptically as composite samples from each replicate (pen) and processed under sterile conditions. Total viable bacterial counts were determined by serial dilution and lawn plating on nutrient agar (incubated at 37°C for 24 h), with results expressed as CFU/g. Fungal counts were determined on Sabouraud dextrose agar (incubated at 25°C for 5 d), with results expressed as CFU/g. The Most Probable Number (MPN) procedure, based on the multiple-tube fermentation was carried out to estimate the coliform count (MPN/g). Lactose broth was applied for presumptive phase using nine tubes (three decimal dilutions of the10% suspension of sample in sterile phosphate buffered saline). Gas production in inverted Durham tube was the indication of possible coliform bacteria growth. After 24 h incubation in 37°C, positive tubes were subcultured on 10 ml brilliant green lactose bile broth with inverted Durham tube. Positive samples were further confirmed using the isolation on MacConkey agar and lactose fermenter isolates were tested for indole production on SIM agar using Kovac’s reagent.

Performance measurements. Body weight (BW) and BW gain were recorded weekly by pen. Feed intake was recorded to calculate feed conversion ratio (FCR; g:g). Mortality was recorded daily and used to calculate livability. The European Production Efficiency Factor (EPEF) was calculated as:

$$\text{EPEF}=\frac{\text{Livability}(\%)\times \text{BW}\left(\text{kg}\right)}{\text{Age}\left(\mathrm{d}\right)\times \text{FCR}}\times 100$$

Statistical analysis. Data were analyzed using R (version 4.5.1). The experimental unit was the pen. The effects of dietary treatments were evaluated using the following general linear model for the completely randomized design (CRD):

$$Y_{ij}=\mu +T_i+{\epsilon }_{ij}$$

where $Y_{ij}$ is the observed response, µ is the overall mean, $T_i$ is the fixed effect of the i th disinfectant treatment, and ${\epsilon }_{ij}$ is the random residual error. Assumptions of normality and homogeneity of variances were verified using the Shapiro–Wilk test and Levene’s test, respectively. For longitudinal data, repeated weekly measures were analyzed by including “week” as a fixed effect in the model. Results are presented as means ± SEM. To enhance precision, exact P-values (e.g., P = 0.003) are reported instead of using only threshold values (e.g., P < 0.05), and percentage changes relative to the control group are included in the results and tables where appropriate.

Results

Microbiological analysis of feed. Total bacterial counts in starter and grower diets were affected by treatment (P < 0.05; Table 9; results expressed as Log₁₀ CFU/g). In starter diets, Nanosil Feed & Air® produced the lowest bacterial counts. In grower diets, formaldehyde treatments (direct or premix) yielded the lowest bacterial counts. Differences among treatments were not significant in finisher diets. Fungal counts were not significantly affected in any phase (P > 0.05; Table 9). Coliform counts did not differ among treatments in starter diets but differed in grower and finisher diets (P < 0.05; Table 9), with the lowest values in formaldehyde-based treatments.

Table 9.

Microbial counts in starter, grower, and finisher diets.

A) Total bacterial counts (Log₁₀ CFU/g) in starter, grower, and finisher diets.
Treatment	Starter	Grower	Finisher
Control	4.89 a	4.62a	4.13
Formalin (feed)	4.37b	3.23b	3.29
Formalin (premix)	4.36b	3.21b	3.34
Termin Gold® (feed)	4.39b	4.13a	4.01
Termin Gold® (premix)	4.51b	3.74c	3.54
Nanosil Feed & Air® (feed)	3.21c	3.82c	3.64
SEM	0.01	0.02	0.03
P-value	<0.001	0.002	0.241
${}^{a--c}$Means within a column without a common superscript differ significantly ($P<0.05$). SEM = standard error of the mean.
B) Total fungal counts (Log₁₀ CFU/g) in starter, grower, and finisher diets.
Treatment	Starter	Grower	Finisher
Control	2.22	3.49	2.95
Formalin (feed)	2.37	2.80	1.92
Formalin (premix)	2.00	3.06	2.88
Termin Gold® (feed)	2.48	3.24	2.07
Termin Gold® (premix)	2.30	3.22	1.70
Nanosil Feed & Air® (feed)	2.18	2.97	1.92
Values represent descriptive $\text{Lo}{\mathrm{g}}_{10}\text{CFU}/\mathrm{g}$from composite dietary samples; no statistical comparison was performed due to single-composite laboratory determinations.
C) Total coliform count ($\mathbf{Lo}{\mathbf{g}}_{10}\mathbf{MPN}/\mathbf{g}$)
Treatment	Starter	Grower	Finisher
Control	3.04	2.88	3.04
Formalin (feed)	3.04	1.10	1.69
Formalin (premix)	3.04	0.85	2.54
Termin Gold® (feed)	3.04	2.58	2.72
Termin Gold® (premix)	3.04	3.04	2.83
Nanosil Feed & Air® (feed)	2.91	1.69	2.73

MPN = most probable number. For samples exceeding the laboratory upper detection limit (>1100 MPN/g), the value of 1100 MPN/g (3.04 Log10 MPN/g) was assigned.

Proximate composition of diets. Disinfectant treatments did not affect crude protein, calculated metabolizable energy, ash, crude fiber, or nitrogen-free extract (P > 0.05; Table 8). Ether extract differed among treatments within phases (P < 0.01), but differences were small and did not indicate a consistent adverse effect.

Table 8.

Proximate composition of experimental diets by feeding phase (%).

A) Starter diet (d 1–10)
Treatment	Moisture		Crude protein	Calculated energy (kcal/kg)	Ash	Crude fiber	Ether extract		Nitrogen-free extract
Control	10.98		23.07	3256	5.53	3.78	4.04		63.57
Termin Gold® (feed)	10.96		22.99	3211	5.44	3.95	3.93		63.69
Nanosil Feed & Air® (feed)	11.02		22.96	3230	5.45	3.72	3.95		63.92
Formalin (feed)	10.97		22.94	3219	5.41	3.67	3.63		64.36
Formalin (premix)	10.94		23.11	3235	5.34	3.77	4.06		63.72
Termin Gold® (premix)	11.03		23.15	3220	5.42	3.84	3.74		63.85
B) Grower diet (d 11–24)
Treatment		Moisture	Crude protein	Calculated energy (kcal/kg)	Ash	Crude fiber	Ether extract	Nitrogen-free extract
Control		11.04	21.31	3324	5.44	3.97	4.30	64.96
Termin Gold® (feed)		10.99	21.33	3326	5.45	3.90	4.22	65.10
Nanosil Feed & Air® (feed)		11.13	21.39	3320	5.43	3.89	4.07	65.22
Formalin (feed)		11.07	21.34	3333	5.49	3.89	4.39	64.88
Formalin (premix)		11.14	21.19	3351	5.34	3.82	4.52	65.12
Termin Gold® (premix)		11.12	21.30	3332	5.32	3.71	4.15	65.52
C) Finisher diet (d 25–35)
Treatment		Moisture	Crude protein	Calculated energy (kcal/kg)	Ash	Crude fiber	Ether extract	Nitrogen-free extract
Control		11.05	19.32	3573	5.42	3.81	5.09	66.36
Termin Gold® (feed)		11.39	19.48	3566	5.32	3.83	4.89	66.49
Nanosil Feed & Air® (feed)		11.18	19.27	3580	5.32	3.72	5.02	66.67
Formalin (feed)		10.91	19.47	3569	5.35	3.95	5.16	66.07
Formalin (premix)		11.20	19.24	3571	5.38	3.69	4.83	66.87
Termin Gold® (premix)		11.04	19.45	3565	5.40	3.85	4.96	66.35

Values represent descriptive proximate analyses from composite dietary samples.

Growth performance. No differences among treatments were detected at d 7 (P > 0.05). From d 21 onward, premix-applied formaldehyde and premix-applied Termin Gold increased BW relative to other treatments (Table 2). BW gain was higher in premix groups during wk 2 to 4 (Table 3). Feed intake was higher in premix groups during wk 2 to 4 (Table 4). FCR differed during later growth (d 28 and 35; Table 5), with lower values in premix formaldehyde and premix Termin Gold. Livability differed at d 21 (Table 6), driven by lower livability in the Nanosil group. EPEF differed among treatments (Table 7), with the highest values in premix groups.

Table 2.

Effects of experimental treatments on body weight (g) of broiler chickens.

Treatment	Day 7	Day 14	Day 21	Day 28	Day 35
Control	149.78	324.87b	477.37b	844.20b	1093.39b
Formalin (feed)	140.81	286.95c	479.09b	867.34b	1134.94b
Formalin (premix)	149.45	355.69a	748.55a	1281.51a	1514.78a
Termin Gold® (feed)	152.63	318.95b	511.43b	844.01b	1081.10b
Termin Gold® (premix)	144.79	350.66a	747.72a	1291.28a	1545.53a
Nanosil Feed & Air® (feed)	144.66	293.83c	489.23b	871.54b	1096.85b
SEM	3.87	8.28	18.47	29.01	22.27
P-value	0.216	<0.001	<0.001	<0.001	<0.001

$a--c$Means within a column with different superscripts differ significantly (P < 0.05). SEM = standard error of the mean.

Table 3.

Effects of experimental treatments on weekly body weight gain (g) of broiler chickens.

Treatment	Week 1	Week 2	Week 3	Week 4	Week 5
Control	114.78	175.10b	152.49b	366.83b	249.19
Formalin (feed)	105.81	146.14c	192.14b	351.25b	265.11
Formalin (premix)	114.45	206.24a	392.86a	532.97a	233.26
Termin Gold® (feed)	117.63	166.32b	192.48b	332.57b	237.09
Termin Gold® (premix)	109.79	205.87a	397.06a	543.56a	254.25
Nanosil Feed & Air® (feed)	109.66	149.18c	195.39b	382.31a	225.31
SEM	3.27	7.75	17.05	33.32	29.8
P-value	0.216	<0.001	<0.001	0.002	0.946

$a--c$Means within a column with different superscripts differ (P < 0.05). SEM = standard error of the mean.

Table 4.

Effects of experimental treatments on weekly feed intake (g/pen) of broiler chickens.

Treatment	Week 1	Week 2	Week 3	Week 4	Week 5
Control	1927.00	5361.00 b	5096.50 b	7975.00 b	8270.50
Formalin (feed)	2069.00	5157.50 b	5224.80 b	8176.30 b	8034.00
Formalin (premix)	2114.00	6789.00 a	10589.50 a	13176.50 a	8414.00
Termin Gold® (feed)	2108.00	5715.50 b	5527.80 b	7264.00 b	7960.00
Termin Gold® (premix)	1878.00	6789.00a	10450.30 a	14292.80 a	8327.80
Nanosil Feed & Air® (feed)	1906.00	5443.00b	5330.50b	7298.30b	7320.50
SEM	69.80	284.76	362.50	784.10	973.30
P-value	0.078	0.001	<0.001	<0.001	0.971

$a,b$Meanswithin a column with different superscripts differ (P < 0.05). SEM = standard error of the mean.

Table 5.

Effects of experimental treatments on feed conversion ratio (FCR; g:g) of broiler chickens.

Treatment	Day 7	Day 14	Day 21	Day 28	Day 35
Control	0.655	1.288b	1.239	1.547a	1.912a
Formalin (feed)	0.735	1.174c	1.266	1.651a	1.775ab
Formalin (premix)	0.708	1.640a	1.365	1.292b	1.838ab
Termin Gold® (feed)	0.694	1.176c	1.273	1.552a	1.863ab
Termin Gold® (premix)	0.649	1.166c	1.333	1.339b	1.690b
Nanosil Feed & Air® (feed)	0.664	1.180c	1.249	1.630a	1.991a
SEM	0.01	0.037	0.017	0.033	0.026
P-value	0.066	0.001	0.221	0.001	0.004

$a--c$Means within a column with different superscripts differ (P < 0.05). SEM = standard error of the mean.

Table 6.

Effects of experimental treatments on livability (%) of broiler chickens.

Treatment	Day 7	Day 14	Day 21	Day 28	Day 35
Control	96.25	98.75	96.25ab	97.50	98.75
Formalin (feed)	100.00	93.75	95.00ab	98.75	95.00
Formalin (premix)	100.00	98.75	100.00a	98.75	100.00
Termin Gold® (feed)	98.75	97.50	96.25ab	97.75	97.50
Termin Gold® (premix)	98.75	98.75	100.00a	100.00	100.00
Nanosil Feed & Air® (feed)	98.75	97.50	87.50b	96.25	96.75
SEM	1.25	1.44	2.67	1.25	1.36
P-value	0.404	0.226	0.018	0.364	0.092

$a,b$Means within a column with different superscripts differ (P < 0.05). SEM = standard error of the mean.

Table 7.

European Production Efficiency Factor (EPEF) of experimental treatments.

Treatment	Final body weight (kg)	FCR	Livability (%)	EPEF
Control	1.093b	1.328	87.5b	205.8c
Formalin (feed)	1.132b	1.320	82.5bc	202.2c
Formalin (premix)	1.515a	1.368	97.5a	308.4b
Termin Gold® (feed)	1.081b	1.312	87.5b	206.1c
Termin Gold® (premix)	1.546a	1.234	97.5a	349.0a
Nanosil Feed & Air® (feed)	1.097b	1.337	78.8c	184.7c
SEM	0.021	0.045	2.30	6.50
P-value	<0.001	0.312	0.008	<0.001

a-c Means within a column with different superscripts differ significantly (P < 0.05). SEM = standard error of the mean. EPEF = (Livability (%) × Body Weight (kg)) / (Age (days) × FCR) × 100.

Discussion

The present study demonstrates that disinfectant efficacy in broiler diets depends on both active ingredient and application strategy. The absence of treatment effects during the first week is biologically plausible because early growth and nutrient absorption are heavily dependent on residual yolk sac utilization and rapid physiological adaptation to solid feed, rendering systemic growth responses less sensitive to minor dietary interventions during this initial period (Leeson and Summers, 2009).

From week 2 onward, the application of disinfectants via premix (specifically for formaldehyde and Termin Gold®) produced superior BW gain and feed efficiency. Mechanistically, applying disinfectants through a premix carrier optimizes particle distribution, improves chemical contact time with feed solids, and prevents localized chemical degradation. This enhanced distribution minimizes pathogen load and suppresses secondary microbial metabolites in the feed, which otherwise trigger mild gut inflammation and depress intake (Avila et al., 2023).

Performance benefits were accompanied by higher feed intake during early growth, which reflects improved feed palatability and reduced subclinical enteritis risks associated with microbial metabolites that can compromise gut mucosal integrity (Stanley and Bajagai, 2022; Stanley et al., 2014). The time-dependent nature of responses is consistent with previous reports showing that formaldehyde–organic acid blends can influence microbial counts and performance across production phases (Sbardella et al., 2015). Improved FCR during later growth supports the concept that lower microbial pressure conserves metabolic energy, allowing absorbed nutrients to be redirected from intestinal immune activation toward somatic tissue deposition (Huyghebaert et al., 2011).

A practical concern with chemical disinfectants is potential interaction with nutrients, especially proteins and vitamins via amino acid cross-linking or lipid oxidation (Leeson and Summers, 2009). In this study, major proximate composition of diets was not adversely affected, suggesting that the inclusion rates and handling conditions were compatible with maintaining expected nutrient specifications. Small differences observed for ether extract were not consistent across phases and did not indicate a reduction in overall diet quality or energy density.

Microbiological results confirmed reduced bacterial contamination, particularly in starter and grower feeds. Silver nanoparticles in Nanosil Feed &Air® possess a high surface-area-to-volume ratio, facilitating rapid binding to bacterial membranes, structural disruption, and generation of reactive oxygen species, producing strong antibacterial effects in starter diets (Seong et al., 2021). However, formaldehyde—especially when applied as a premix—was most effective in grower diets, which aligns with its ability to achieve uniform distribution within the feed matrix and alkylate microbial functional proteins and nucleic acids, providing sustained broad-spectrum sanitation (Ricke et al., 2019). The lack of significant effects on fungal counts likely reflects the dominant role of moisture, free water activity ($a_w$), and storage conditions in fungal proliferation; therefore, fungal control may require integrated management beyond chemical disinfectant inclusion (Kabak et al., 2006).

Overall, the findings support premix application as an effective strategy for formaldehyde-based disinfectants, improving feed hygienic quality and broiler performance without compromising proximate nutrient composition. Future studies should evaluate disinfectant stability under different storage durations and environmental conditions and examine impacts on gut microbiota and welfare outcomes (Ricke et al., 2019; Omidi et al., 2025).

Conclusion

In conclusion, premix-applied formaldehyde-based disinfectants effectively improve broiler growth performance, feed intake, and feed conversion efficiency by mitigating dietary bacterial contamination, without compromising the proximate nutrient composition of the feed.

A primary limitation of this study is that microbial stability and disinfectant efficacy were evaluated under controlled experimental conditions over a single production cycle, without evaluating long-term feed storage effects or the impacts on intestinal histomorphology and microbiome dynamics. Future research should investigate the long-term stability of premixed disinfectants across diverse ambient storage environments and elucidate their specific effects on gut health and broiler welfare.

Author contributions

Amir Janfadah: conceptualized the initial idea, planned the study design, prepared the animal facility, collected the samples, curated the data from the birds, assessed and collected the microbiological and feed analytical data, analyzed the data, performed the initial statistics, and wrote and approved the initial draft.

Seyed Ahmad Madani: conceptualized the initial idea, planned the study design, revised the details of the design, supervised the data acquisition and analysis, validated the methodology and data, interpreted the results, co-wrote the initial draft, revised and approved the final manuscript.

Mohammad Reza Mohammadian: planned the study design regarding the feed microbiological methods and added more details to the design, performed the microbiological methods, collected and analyzed the microbiological data, approved and revised the initial draft, and revised and approved the final manuscript.

Seyed Mohammad Mahdi Hashemian: co-planned the detailed study design, supervised the data collection from the poultry house, investigated the animals’ rearing and performance data collection, performed and analyzed the feed analysis, revised and approved the final manuscript.

Funding

This research was funded by Faculty of Veterinary Medicine, University of Tehran.

Declaration of AI and AI-assisted technologies in the writing process

During the preparation of this work the authors used GapGPT (GPT-5.6) for literary and grammatical editing of the article text to improve the readability. After using this service, the authors reviewed and edited the content as needed and take full responsibility for the content of the publication.

Disclosures

The authors declare no conflict of interest.