IMPACT OF BORON AND NANO-BORON ON THE HETEROPHIL ... · Dhekra Mahdi Mousa, Balqees Hassan Ali ** Department of pathology and poultry disease, College of Veterinary Medicine, University

Basrah Journal of Veterinary Research,Vol.17, No.3,2018 Proceeding of 6th International Scientific Conference,College of Veterinary Medicine University of Basrah,Iraq

290

IMPACT OF BORON AND NANO-BORON ON THE

HETEROPHIL /LYMPHOCYTE RATIO AND

HISTOPATHOLOGICAL CHANGES OF LIVER AND KIDNEY

IN BROILER CHICKENS INFECTED WITH Escherichia coli

Dhekra Mahdi Mousa, Balqees Hassan Ali

** Department of pathology and poultry disease, College of Veterinary Medicine,

University of Baghdad.

Keywords: Boron, Escherichia coli, Chickens.

Corresponding author: [email protected]

ABSTRACT

The aim of this study was to evaluate the effect of boron and nano-boron on

heterophil/lymphocyte ratio, the histopathological changes of liver and kidney in

broiler chicks and bird resistance against E.coli. A total of two hundred and forty Ross

broiler chicks at one day old were used and distributed randomly to eight equal groups

and were treated from one day to the end of the experiment as follows: B10 was

received boron 10mg/L of drinking water, B20 boron 20mg/L, B40 boron 40mg/L,

NB10 was received nano-boron 10mg/L, NB20 nano-boron 20mg/L, NB40 nano-

boron 40mg/L, either control groups: C+ was vaccinated only, C- not treated pre

challenge. The maternal immunity was measured in the 2nd day for all groups by

ELISA test; all groups were vaccinated (except group C-) with Newcastle disease

vaccine (LaSota) at (15and25days old) and infectious bursal disease vaccine at

(16days old) via drenching route. Crystalline structure and size of nano-boron powder

was determined by X-ray diffraction analyses. Blood samples were collected at

35days old through the jugular vein. The livers and kidneys were removed and tissue

sections were stained with hematoxylin and eosin stain for histological examination at

35and after challenge for all grroups at 40days old. The results showed as follows:

boron group B10 recorded significant decrease in heterophil/lymphocyte ratio than

other doses of boron but the higher heterophil/lymphocyte ratio was recorded in nano-

boron doses in comparison with control groups C+ and C-. Histological examination

results pre challenge test with E.coli referred a pathological changes in the liver and

kidney tissues due to boron and nano-boron in all groups like aggregation of


291

mononuclear cells, vacuolar degeneration, proliferation of kupffer cells and

granulomatous lesions. After challenge test at 35days old, the percentage of morbidity

rate was 100% in control groups C+, C- and 40% recorded in group B40 while other

groups (B10, B20, NB10, NB20 and NB40) recorded 60%, mortality rate was zero

during the trial period, at 40days old the results showed histopathological changes in

the liver and kidney in all groups, these changes were more severe in control groups

like (sever vacuolation either hydropic or fatty degeneration). In conclusion, the

results of present study indicated that the addition of boron and nano-boron to

drinking water reduced the stress factor and increase the resistance of birds against

E.coli infection.

INTRODUCTION

Boron has a physiological amount effects usefully on substantial number of

metabolic processes in humans and animals [1] which include activity of many

metabolic enzymes, as well as the metabolism of steroid hormones and several

micronutrients, including calcium, magnesium, and vitamin D. Growing evidence has

demonstrated that boron plays a central role in the development of animals [2]. The

antibacterial properties of boron-containing compounds are well known [3] and it is

mostly found in the form of boric acid or borate [4], thus boric acid is mildly

antimicrobial [5]. Given the established biological effects of boron in the body,

namely as antioxidant [6], influencing energy metabolism [7] and mineral formation

of bones and cartilages [8] can be considered that boron meets the requirements to be

considered an essential trace element in poultry [9].

Nanoparticles (NPs) can be used as a supplemental source of trace minerals in

diets [10]. So when the active ingredient is nanostructured, the substance stability

increases due to protection against oxidant agents, enzymes and other components

[11]. Additionally, their small size makes it possible to surpass physiological barriers

and characteristics such as high surface area to volume ratio, which may allow for

increased interaction with pathogen membranes and cell walls [12], therefore

nanotechnology and nanobased therapeutics may entirely change the whole concept of

healthcare in coming years [13]. There are limited studies available on the pure boron

NPs [3].

Avian pathogenic Escherichia coli (APEC) are caused a serious disease in

poultry [14]. Although antibiotic agents have been used to control APEC infections


292

[15], E.coli has become one of the microorganisms that are generally resistant to

antimicrobials [16]. Therefore, this study aimed to study the effect of boron and nano-

boron and enhanced the resistance of broiler chicks against E.coli infection.

MATERIALS AND METHODS

A total of two hundred and forty, 1 day old broiler chicks, were weighted and

randomly divided into 8 groups (each group 30), boron groups (B) 10, 20 and 40 mg

boron/L of drinking water, nano-boron groups (NB) 10, 20 and 40 mg nano-boron/L

and two control groups C+ and C-, group C+ was vaccinated only, group C- not

treated. Maternal immunity was determined in the 2nd day for vicinal program; all

groups were vaccinated except group C- against ND and IBD virus. The experimental

period lasted for 40days. Boric acid was used as the boron source, the boron NPs was

imported from China (Hefei EV NANO Technology Corporation, Ltd- china), some

of them were sent to the central service laboratory at the Faculty of Education for

Pure Sciences/ Ibn Al-Haytham to determine the crystalline structure of the nano-

boron by dropping the X-ray spectra (XRD 6000) on the examined material [3].

Figure 1 shows the X-ray diffraction patterns of nano-boron, the diffraction peaks at 2

θ (degrees) of 19.96°, 28.80°, 36.79°, 44.95°, 52.32°, 64.34° and 78.32°, but the other

degrees in the figure is a impurities of silicon carbide. All these peaks indicated as

cubic structure of the lattice nano-boron. The broad in the diffraction peaks reflects

the small size of the crystalline size. The size of nanoparticles was estimated by

DebyScherrer, s equation [17]:

L=

Where it symbolizes (λ =0.167 A°) it’s the wave length of X-ray, β is the peak width

of half maximum (FWHM) in radian which have (0.33480), θ is angle diffraction of

Bragg,s in degree, the crystalline size of boron NPs found (20.6) nanometer.

Ultrasonic device was used to break the aggregation of particles [18] and homogenize

the mixture. Blood samples 80were collected at 35days old through the jugular vein

[19]. Five chickens of each group were selected at 35days old, and E.coli (2×108)

CFU/ml infection [20] was made in respiratory and digestive tract. Morbidity and

mortality rate were recorded. At 35and after challenge test at 40days old, liver and


293

kidney sections were collected and fixed in 10%formalin solution for fixation, then

dehydrated by gradedethanol, cleared in xylene and embedded in paraffin,sectioned to

five microns and stained with hematoxyline and eosin [21]. The Statistical Analysis

System- SAS [22] program was used to study the effect of different factors in study

parameters.

Figure 1 The X-ray diffraction patterns of nano-boron

RESULTS

Hetrophil/Lymphocyte Ratio (H/L)

Results in table 1 revealed there were a significant difference (P 0.01) between

C+ and all groups. Comparing mean H/L ratio between all the groups showed that

group B10 was the lower level followed by NB20, B40, B20, NB40, NB10 and C-

while C+ showed the higher H/L ratio.

Morbidity and Mortality Rate after Challenge Test

Morbidity rate was as follows in (table 2) depending on the cases of diarrhea and

rales sound as followed: B10, B20, NB10, NB20, NB40 (3 cases), B40 (2 cases), C+,

C- (5 cases). The results showed significant differences between B40 with all the

groups, also there was no death reported in infected chicken.

Histopathology before Challenge Test

In group B10 the histopathological manifestation of liver showed multifocal

mononuclear cells (MNCs) infiltration in liver tissue mainly in portal area and around

dilated central vein, also showed periductal MNCs aggregation accompanied with

mild ductal dilation, while other section revealed moderate aggregation of lymphocyte

(fig A) in liver parenchyma with sinusoid dilation together with prominence of

kupffer cells. In group B20 showed main hepatic findings were characterized by

variable degree of perivascular MNCs aggregation consist mainly of lymphocyte and


294

macrophage, also the kupffer cells that lining the sinusoids were proliferated with no

clear pathological alteration in hepatic tissue. In group B40 showed moderate portal

enlargement was observed due to periductal fibrosis. The predominant hepatic lesion

characterized by presence of small granulomatous lesion mainly in portal area (fig B).

Also presence small granulomatous foci surrounded with mild fibrosis zone in portal

area. In group NB10 showed that the hepatic tissue containing multiple MNCs

aggregation (fig C) mainly around blood vessels and portal region, also perivacuolar

MNCs aggregation accompanied with mild degeneration of hepatic lobules. In group

NB20 showed that liver section exhibited similar microscopic observation to previous

group that showed intense MNCs aggregation mainly around central vein (C.V)

together with kupffer cell proliferation (fig D), also presence of inflammatory cells in

other dilated congested blood vessels. In group NB40 showed that the majority of

hepatic blood vessels containing large number of inflammatory cells and evidence of

sinusoidal congestion with organized hepatic cords architecture together with kupffer

cells proliferation. Also exhibited portal tract enlargement due to vasodilation and

moderate ductal dilation accompanied with MNCs infiltration consist mainly of

lymphocyte and macrophage (fig E). In group C+ the microscopic lesion of liver

sections showed dilation and congestion of sinusoids with focal MNCs aggregation in

the hepatic parenchyma, other section showed large cellular aggregation consist

mainly of polymorphonuclear cells seen mainly in portal area (fig F), focal interstitial

polymorphonuclear cells infiltration with slight septal fibrosis mainly adjacent to the

portal area, also moderate vacuolar degeneration were noticed in liver parenchyma

mainly in portal area with evidence of vacuolar congestion.

In kidney, group B10 showed moderate focal interstitial MNCs infiltration

consist mainly of lymphocytes, also mild thickening of renal capsule with

inflammatory cellular infiltration seen in other section accompanied with severe

tubular degeneration (fig G) and vacuolar degeneration of some tubules with mild

interstitial congestion. In group B20 showed mild histopathological changes were

observed include tubular dilation, flat epithelial lining and interstitial hemorrhage (fig

H). In group B40 no clear histopathological changes were recorded except little

number of cortical cysts was observed in renal tissue. In group NB10 showed tubular

epithelial lining were swollen and hydropic with mild perivascular MNCs aggregation

associated with focal interstitial MNCs infiltration and slight vacuolar congestion.

Also prominence of basophilic cortical tubules was observed in other section

accompanied with moderate cellular infiltration (fig I). In group NB20 showed focal

MNCs infiltration were recorded in renal parenchyma associated with acute cellular

swelling of the major renal tubules (fig J), severe cellular swelling of renal tubules

and mild to moderate interstitial fibrosis that divided renal tissue in multiple lobules.

In group NB40 showed moderate tubular dilation with focal interstitial MNCs

infiltration was observed in renal tissue (fig K), also congestion of interstitial tissue

with slight tubular congestion occurred in other section. In group C+ showed

moderate to severe congestion of kidney interstitial tissue associated either with

cellular swelling or atrophy of adjacent renal tubules (fig L), other renal observation


295

showed glomerular hypercellularity with severe narrowing of bowman space. Also

there was vacuolar degeneration of tubular epithelium accompanied with appearance

of basophilic tubules mainly in cortical area. In group C- showed no clear

pathological changes have been detected in both liver and kidney tissues with normal

structural details.

Histopathology after Challenge

In group B10 the principle hepatic lesion was manifested by intensive

perivascular aggregation consist of macrophage and lymphocyte seen mainly around

C.V with no alterations in liver parenchyma, together with evidence of occlusion of

portal vein by eosinophilic edematous fluid. In group B20 the majority of hepatic

blood vessels were congested and containing large number of MNCs consist mainly

of lymphocyte and macrophage, also marked dilation of portal vein with severe

periductal fibrosis, in other section hyperplastic epithelium and cellular debris in

ductal lumen. In group B40 the pathognomic lesion characterized by massive

aggregation of leukocytes around hepatic blood vessels and hepatic parenchyma

mainly consist of lymphocytes and macrophage together with prominence of kupffer

cells. In group NB10 moderate to severe histopathological change were observed in

both hepatic and renal tissues of this group, the majority of liver section appears

multifocal MNCs aggregation consist mainly of macrophage and lymphocyte seen

either in dilated sinusoids or around congested blood vessel. Other manifestation

showed biliary epithelial hyperplastic accompanied with medial muscular hypertrophy

of hepatic artery, portal MNCs infiltration and marked venous dilation was recorded

mainly in portal tract accompanied with focal lymphocytic accumulation around

reminant of bacterial colonies (fig M). In group NB20 small granuloma was observed

in liver tissue consist of MNCs. In group NB40 showed similar observation to

previous group, characterized by granulomatous reaction and proliferation of kupffer

cells together with marked perivascular MNCs infiltration. In group C+ showed

granulomatous lesions consist of MNCs (fig N) with evidence of severe vaculation of

surrounding parenchyma, also the portal tract exhibited fibromuscular hypertrophy of

portal blood vessels (hepatic artery and portal vein) associated with individual liver

cell necrosis. While other section showed focal polymorphonuclear leukocytes

aggregation around small ductules causing disarrangement of liver parenchyma. In

group C- showed focal polymorphonuclear cells infiltration in liver parenchyma and


296

severe vacuolar degeneration of liver cells (fig O), also disruption of hepatic cord

(with evidence of individual cell necrosis may reported).

In group B10 kidney showed moderate focal interstitial MNCs aggregation with

moderate vacular degeneration of surrounding tubules, together with evidence of

cortical hemorrhage and basophilic cortical tubular presence. In group B20 main renal

lesion showed focal lymphocytic infiltration together with tubular epithelial swelling,

marked dilation and congestion of renal blood vessels. In group B40 the renal section

expressed similar lesion to previous group in which focal lymphocytic aggregation

was occurred with variable degrees of tubular epithelial degeneration, as well as

tubular atrophy with infiltration of neutrophils and MNCs in the interstitial tissue

(figure P) and vacuolation of epithelial lining renal tubules with interstitial

haemorrhage. In group NB10 multifocal interstitial MNCs infiltration noticed with

moderate tubular dilation, as well as cortical cyst manifestation with moderate tubular

epithelial vaculation. In group NB20 both tubular and glomerular tissues showed no

clear microscopical changes, while other section revealed focal MNCs infiltration

with interstitial congestion associated with fibromuscular hypertrophy of renal blood

vessel. In group NB40 showed that renal parenchyma containing large number of

regenerated basophilic tubules together with mild tubular degeneration of adjacent

tubules. In group C+ showed moderate vacuolar epithelial degeneration of many renal

tubules either hydropic or fatty degeneration (fig Q) and moderate to severe cystic

tubular dilation accompanied with fatty degeneration of epithelial lining of

surrounding tubules. Other renal observation showed mild cellular infiltration seen in

the interstitial tissue. In group C- showed that the epithelial lining of many renal

tubules were either hydropic swollen or necrotic associated with sever degeneration of

glomerular tuft (fig R), also focal poly MNCs infiltration was noticed, in other section

forming granulomatous like lesion, evidence of interstitial hemorrhage with variable

degree of tubular dilation.


297

Table 1 Hetrophil / Lymphocyte ratio of different groups at 35 days old of

chicken (Mean± SE)

The group Mean ± SE

B10 5.92 ± 0.36 d

B20 9.35 ± 0.81 bc

B40 8.61 ± 0.64 c

NB10 10.77 ± 0.83 bc

NB20 8.18 ± 0.59 c

NB40 10.62 ± 0.74 bc

C+ 13.28 ± 0.82 a

C- 11.96 ± 0.62 bc

Level of sig. **

** (P≤0.01).

Means having with the different letters in same column differed significantly.

Table 2 Morbidity rate (%) of different groups after challenge test (E.coli) at 40

days old

Group Morbidity (%)

B10 60.00 b (3)

B20 60.00 b (3)

B40 40.00 c (2)

NB10 60.00 b (3)

NB20 60.00 b (3)

NB40 60.00 b (3)

C+ 100.0 a (5)

C- 100.0 a (5)

Level of sig. **

** (P≤0.01) N=40

Means having with the different letters in same column differed significantly.


298

Figure A: histopathological section in liver (B10) moderate aggregation of

lymphocyte in parenchyma with sinusoid dilation together with prominence of

kupffer cells (H&E stain X40). Figure B: liver (B40) presence of small

granulomatous lesion mainly in portal area (H&E stain X10). Figure C: liver

(NB10) the hepatic tissue containing multiple MNCs aggregation (H&E stain

X20). Figure D: liver (NB20) intense MNCs aggregation mainly around central vein

together with kupffer cell proliferation (H&E stain X40). Figure E: liver (NB40)

portal tract enlargement due to vasodilation and moderate ductal dilation

accompanied with MNCs infiltration consists mainly of lymphocyte and macrophage

(H&E stain X10). Figure F: liver (C+) large cellular aggregation consists mainly of

poly morphic nuclear cells seen mainly in portal area (H&E stain X20).

A B

C

F E

C

D


299

Figure G: histopathological section in kidney (B10) mild thickening of renal capsule

with inflammatory cellular infiltration seen accompanied with severe tubular

degeneration (H&E stain X20). Figure H: kidney (B20) flat epithelial lining of renal

tubules with interstitial hemorrhage (H&E stain X20). Figure I: kidney (NB10)

prominence of basophilic cortical tubules was observed accompanied with moderate

cellular infiltration (H&E stain X20). Figure J: kidney (NB20) focal MNCs

infiltration was recorded in renal parenchyma associated with acute cellular swelling

of the major renal tubules (H&E stain X40). Figure K: kidney (NB40) focal

interstitial MNCs infiltration (H&E stain X20). Figure L: kidney (C+) moderate

congestion of blood vessels in the interstitial tissue associated either with cellular

swelling or atrophy of adjacent renal tubules (H&E stain X20).

H

I J

k L

G


300

M N

O P


301

Figure M: histopathological section in liver (NB10) marked venous dilation was

recorded mainly in portal tract accompanied with focal lymphocytic accumulation

around remnant of bacterial colonies (H&E stain X10). Figure N: liver (C+)

granulomatous lesions consist of poly MNCs in portal region (H&E stain X10).

Figure O: liver (C-) severe vacuolar degeneration of liver cells (H&E stain X4).

Figure P: kidney (B40) tubular atrophy with infiltration of neutrophils and MNCs in

the interstitial tissue (H&E stain X20). Figure Q: kidney (C+) moderate vacuolar

epithelial degeneration of many renal tubules either hydropic or fatty degeneration

(H&E stain X40). Figure R: kidney (C-) the epithelial lining of many renal tubules

were either hydropic swollen or necrotic associated with sever degeneration of

glomerular tuft (H&E stain X40).

DISCUSSION Hetrophil/Lymphocyte ratio (H/L) `

Group C+ recorded higher mean H/L ratio followed by others groups expect the

B10 group was recorded the lower mean H/L ratio. Several studies have indicated the

effect of stress factors on the proportion of hetrophil to lymphocyte [23]. According to

the results, boron and nano-boron are considered an antistress due to decreasing

heterophil numbers to lymphocyte. In research by [24] who found that the decrease in

the number of heterophil may be due to role of boron in improving the immune

response in the body. In study by [25] stated that the number of lymphocytes was high

in comparison to the low number of heterophil when using boron due to role of boron

in enhance of immune system. The regulatory role of boron in the metabolism and

respiratory mechanism impacted the immune organs and lymphocytes [26]. As for

control group C+ observed height in H/L ratio agree with [27] whom reported that

stress from vaccines led to increased secretion of corticosteroid which produced from

the cortex of adrenal glands in response to various stress factors which leads to the

Q R


302

death of lymphocytes and then leads to reduced their numbers compared to numbers

of heterophil cells.

Morbidity and mortality rate after challenge test

The lowest morbidity rate 40% was in group B40 while no significant

differences between treated groups in comparison with C+ and C- which recorded

highest morbidity rate 100%. Indicting an improved immune response by adding

boron and nano-boron compared to control groups, these results agree with [28]

explained the role of boron in improvement the cellular and humeral immune

response and immune function by enhance Fc receptor expression, interleukin-6

production, production of cytokines by increasing the production of tumor necrosis

factor and interferon- γ after a stress or disease challenge. In addition, mortality rate

was zero in all groups.

Histopathology before and after Challenge

The results of histological examination of the liver in all boron groups before

experimental infection showed that boron consumption led to tissue pathological

changes, the results agreement with [29] who study the effects of drinking boron on

liver tissue of Africa ostrich chicks and found that addition of boron in drinking water

adversely affected the development of Africa ostrich chicks' livers and caused obvious

pathological changes. For kidney the results of histological examination in all boron

groups before experimental infection showed that boron consumption led to tissue

pathological changes, so the results agreement with [30] who found that addition of

boron in drinking water caused obviously adverse effect on kidney tissue in ostrich

chicks which including characters of cellular swelling and dissociation, loose

cytoplasm, nuclear pyknosis, basement membrane detachment in some tubular cells,

debris deposit was accumulated in the tubular lumen, tubular dilatation and

degeneration, interstitial inflammation and cast formation in most of the lumens. As

for the results of histological examination of the liver and kidney in all nano-boron

groups before experimental infection were showed pathological changes due to nano-

boron consumption. Since nano-boron is first used in chickens, there is no research

about it. The results of histological examination of the liver and kidney in all boron

groups after E.coli challenge led to pathological changes in addition to previous

lesions, the results agreement with [31] who found pathological changes in liver after

infected intra-tracheally with avian pathogenic E.coli (APEC) were seen intra-and


303

perivascular and parenchymal lymphocyte infiltrations in tissue. In kidney showed the

histopathological manifestation in addition to previous lesions, the results agreement

with previous search who found nuclear pyknosis in tubular epithelium and

enlargement of Bowman’s space. In group C- showed the histopathological

manifestation of liver and kidney. The results after E.coli challenge showed that

histopathological changes were the strongest in groups C+ and C- due to role of boron

and nanoboron in antioxidant defense mechanisms, so the results agreement with [32]

who found that addition boron in rations of Bandarah chickens improved antioxidants

enzymes total antioxidant capacity and superoxide dismutase (TAC and SOD) and

significantly decreased in plasma lipid peroxidation based on malondialdehyde

(MDA) levels. In study by [33] explained the researchers hypothesis of boron’s anti-

inflammatory effects result from inhibition of the oxidative burst by scavenging cells

macrophage and excessive activity by hetrophils, also boron boosts free radical

scavenging by raising levels of a triumvirate of antioxidant enzymes in blood and

cells: SOD, catalase and glutathione peroxidase; enhanced antioxidant defense

mechanisms and regenerated damaged liver, kidney, and brain tissues [34]. As for

nanoboron, the results indicated to role of nanoboron in the tissues of liver and kidney

as antioxidant agent. In conclusion, the results of present study indicated that boron

and nano-boron consumption in drinking water reduced the stress factor and increase

the resistance of birds against E.coli infection.

REFERENCES

1. Hunt, C.D. and Idso, J.P. (1999). Dietary Boron as a physiological regulator of

the normal inflammatory response: a review and current research progress. J

Trace Elem Exp Med; 12:221-233.

2. Hunt, C.D. (2012). Dietary boron: progress in establishing essential roles in

human physiology. J Trace Elem Med Bio, 26: 157-160.

3. Akbar, W.; Karagoz, A.; Basim, G.; Noor, M.; Syed, T.; Lum, J. and

Unluagac, M. (2015). Nano-boron as an Antibacterial Agent for

Functionalized Textile. MRS Proceedings, 1793: 53-57.

4. Dean, J.A. (1999). Lange's handbook of Chemistry 15th edition. McGraw-Hill

New York. Cytokines and Bone Metabolism. M Gowen Boca Raton, CRC

Press: 299–324.


304

5. Irschik, H.; Schummer, D.; Gerth, K.; Höfle, G. and Reichenbach, H. (1995).

“The tartrolons, new boron-containing antibiotics from a myxobacterium,

Sorangium cellulosum". The Journal of antibiotics. 48 (1): 26–30.

6. Janeway, C.A.; Travers, P.; Walport, M. and Shlomchik, M.J. (2005).

Immunobiology: The Immune System, In Health and Disease, 6th edition.

New York: Garland Science.

7. Bakken, N.A. and Hunt, D.C. (2003). Dietary Boron decreases peak pancreatic

in situ insulin release in chicks and plasma insulin concentrations in rats

regardless of Vitamin D or Magnesium status, The Journal of Nutrition, 133:

3577-3583.

8. Hunt, C.D. (1989). Dietary boron modified the effects of magnesium and

molybdenum on mineral metabolism in the cholecalciferol deficient chick.

Biol. Trace Elem. Res. 22: 201-220.

9. Lavinia, S.; Dan, D.; Sandu, S.D.; Ioan, P.; Elena, P. and Eliza, S. (2014).

Effects of new mineral sources for boron supplementation on meat quality in

broilers. Rom Biotechnol Lett 19(4): 9585–9596. 26.

10. Mohapatra, P.; Swain,R.K.; Mishra, S.K.; Behera, T.; Swain, P.;

Behura,N.C.; Sahoo, G.; Sethy, K.; Bhol, B. P. and Dhama, K. (2014). Effects

of Dietary Nano- Selenium Supplementation on the Performance of Layer

Grower Birds. Asian Journal of Animal and Veterinary Advances, 9: 641-652.

11. Brandão, H.M.; Gern, J.C.; Vicentini, N.M.; Pereira, M.M. and Andrade,

P.V.D. (2011). Nanotecnologia: a próxima revolução na agropecuária. Rev.

CFMV, ano XVII (53): 61-67.

12. Blecher, K.; Nasir, A. and Friedman, A. (2011). The growing role of

nanotechnology in combating infectious disease. Virulence, 2: 395–401.

13. Chakravarthi, V.P. and Balaji, N.S. (2010). Applications of nanotechnology in

veterinary medicine. Vet. World, 3: 477-480.

14. Barnes, H.J., Nolan, L.K., and Vaillancourt, J.P. (2008). Colibacillosis. In:

Saif Y.M.; Fadly, A.M.; Glisson J.R.; McDougald, L.R.; Nolan, L.K.;

Swayne, D.E.; eds. Disease of poultry, 12th edn. Ames, Iowa, USA: Blackwell

Publishing Professional, pp. 691–737.

15. Gregersen, R.H.; Christensen, H.; Ewers, C. and Bisgaard, M. (2010). Impact

of Escherichia coli vaccine on parent stock mortality, first week mortality of


305

broilers and population diversity of E. coli in vaccinated flocks. Avian Pathol

39: 287–95.

16. Zhao, S.; Blickenstaff, K.; Bodeis-Jones S.; Gaines S.A.; Tong E. and

McDermott P. F. (2012) Comparison of the prevalences and antimicrobial

resistances of Escherichia coli isolates from different retail meats in the United

States. Applied and Environmental Microbiology. 78(6): 1701–1707.

17. Patterson, A. (1939). "The Scherrer Formula for X-Ray Particle Size

Determination". Phys. Rev. 56 (10): 978–982.

18. Mahbubul, I.M.; Chong, T.H.; Khaleduzzaman, S.S.; Shahrul, I.M.; Saidur,

R.; Long, B. D.; Amalina, M. A. (2014). Effect of Ultrasonication Duration on

Colloidal Structure and Viscosity of Alumina−Water Nanofluid. Industrial and

Engineering Chemistry Research. 53: 6677-6684.

19. Allan, W.H.; Lancaster, J.E. and Toth, B. (1978). Newcastle Disease

Vaccines, their production and use. Food and Agricultural Organization of the

United Nations, Rome, Italy.

20. Sambrook, J.; Fritsch, E. and Maniatis, T. (1989). Molecular Cloning.

Alaboratory Manual. New York. Cold Spring Harbor Laboratory Press, USA.

21. Thornton, D. H. and Pattison, M. (1975). Comparison of vaccines against

infectious bursal disease. J. Comp.Path. 85: 597-610.

22. SAS. (2012). Statistical Analysis System, User's Guide. Statistical. Version

9.1th ed. SAS. Inst. Inc. Cary. N.C. USA.

23. Kassab, A.; AL-senied, A. and Injidi, M.H. (1992). Effect of dietary ascorbic

acid on the physiology and performance of heat stressed broilers. Proceedings

of the 2nd symposiam. Ascorbic acid in domestic animals Ittingen,

Switzerland. PP. 270-285.

24. Basoglu, A.; Baspinar, N. Ozturk A.S. and Alkalin, P.P. (2010). Effects of

boron administration on hepatic steatosis, hematological and biochemical

profifi les in obese rabbits. Trace Elements and Electrolytes 27: 225 – 231.

25. Hunt, C.D. (2003). Dietary boron: an overview of the evidence for its role in

immune function. J Trace Elem Exp Med, 16: 291-306.

26. Devirian, T. A. and Volpe, S. L. (2003). The physiological effects of dietary

boron. Crit. Rev. Food Sci., 43: 219-231.


306

27. Compton, M. M.; Gibbs, P. S. and Johnson, L. R. (1990). Glucocorticoid

activation of deoxyribonuclric acid degradation in bursal lymphocytes. Poult.

Sci. 69: 1292-1298.

28. Armstrong, T. A.; Spears J. W. and Lloyd, K. E. (2001). Inflammatory

response, growth and thyroid hormone concentrations are affected by long –

term boron supplementation in gilts. J. of Anim. Sci. 79: 1549 – 1556.

29. Wang, W.; Peng, K.; Xiao, K.; Zheng, X.; Zhu, D.; Yang, Z.; Tang, J.; Sun, P.

and Wang, J. (2014). Effect of Drinking Boron on Liver Tissue of Africa

Ostrich Chicks. Vol.41. Issue (9): 136-139.

30. Wang, J.; Zhong, J.M.; Sun, P.P.; Xiao, K.; Tang, J.; Wang, W. and Peng,

K.M. (2015) Effect of boron administration on the morphology of ostrich

chick kidney tissue. Pak Vet J, 35(4): 489-493.

31. Anta, O.E., Susanne Glodde +9 authors (2008). The chicken as a natural model

for extraintestinal infections caused by avian pathogenic Escherichia coli

(APEC), Microbial Pathogenesis, 1–9.

32. El-Saadany Amina, S.; EL-Barbary Amal, M. and Shreif Effat, Y. (2016).

The influence of dietary boron supplementation on performance and some

physiological parameters in bandarah chickens 1- Growing period. Egypt.

Poult. Sci. Journal (36) (IV): (1131-1146).

33. Scorei, R.I.; Ciofrangeanu, C. and Ion, R. (2010). In vitro effects of calcium

fructoborate upon production of inflammatory mediators by LPS-stimulated

RAW 264.7 macrophages. Biol Trace Elem Res. 135(1-3): 334-344.

34. Lara Pizzorno, M.A. MDiv, L.M. (2015). Nothing Boring About

Boron.Integrative M Pizzorno-Boron edicine. 14(4): 35–48.

Documents

IMPACT OF BORON AND NANO-BORON ON THE HETEROPHIL ... · Dhekra Mahdi Mousa, Balqees Hassan Ali ** Department of pathology and poultry disease, College of Veterinary Medicine, University