INTRODUCTION:

Brucellar spondylitis is without a doubt the most widely recognized confinement of dynamic brucellosis.¹ There are several serious general medical issue which are dominion in developing countries, one of the most common that classified as zoonosis with a worldwide distribution is Brucellosis.² Controlling brucellosis is considered a big issues especially in developing countries, due to the inconceivability of totally killing it in zoonotic source, In Iraq the distribution of brucellosis has expanded in close period of time.

Some Zoonotic diseases like brucellosis can developed with complications including numerous organs, foremost is musculoskeletal system and osteoarticular complications and their particular sites of involvements.³ The disease can create many problems in the biological system. Excretion of ions which is related to oxidative stress like (NO-2, NO-3) have a real function in physiological system like it is involvement in lipid peroxidation, protein deactivation and DNA damaging. Additionally, brucellosis affect on the body’s antioxidant defense frame work through prolonged of oxidative stress status.^4,5 Oxidative stretch coming about from infections can cause modification within the serum micronutrient (elements)⁶. Trace elements level could alliterated during infected with Brucellosis⁷. Trace elements classified as a micronutrient, Which were present in macro algae, Abutilon hirtum^8,9, these micronutrients play a vital role in catabolism and anabolism reactions, some of the elements are involved in biochemical reactions as a co-factor for activation of enzymes reactions. The alteration in their concentration is affect the biological system^10,11. Fe considered as an important micronutrient that plays a central role in cell growth processing, proliferation as well as to cellular metabolism. So the presence of any inflammation or neoplasia can cause Fe metabolism imbalanced¹². Zn has several important functions in a biological system, It is considered as an antioxidant, Also it is involved in some enzymes function, like polymerase enzymes¹³. Cu it is an element that has several biological functions it plays a role in superoxide dismutase enzymes as well as to its antioxidant properties⁴. Selenium has essential biological functions because of it is involvement in selenoproteins, it is also considered as a cofactor for Glutathione peroxidase GPX¹⁵. Aluminium is not known to involved in biological processes, and it is not required by the biological system¹⁶. Other minerals that warrant attention are silicon and chromium, silicon has been reported as important for bone formation, Also researchers concluded that silicon make the bone matrix more classifiable because it has an effect on collagen¹⁷. Chromium is a metal present in water, soil, and biological systems, chromium to be an essential element, it is involved in impeded glucose tolerance, the analysts distinguished Cr as an indispensably portion of the protein known as glucose tolerance factor (GTF). Cr also involved in carbohydrates metabolism as well as to metabolism of proteins and lipids mainly by increasing the efficiency of insulin¹⁸. Elemental composition in herbals, like Hyperichi herba,consedered as a herbal remedy¹⁹. Some elements become toxic when an increase from the normal level^20,21. Elements levels were related to type of personal work as proved by several reviewers²³. The present study focused on disparities in some metals level based on ether the pathogenesis of the disease or as a result of infection.

MATERIALS AND METHODS:

Study population:

Samples have been collected from outpatient clinics in Serdem private hospital and Erbil international hospital in Erbil, Iraq at the period from June 2018 to August 2019. The patients were diagnosed by an orthopedic and spine surgeons based on their clinical manifestations, serology tests, X-ray and MRI. The researchers followed the volunteer basis role when they collected the samples, and the study proposal was reviewed and proved by the ethical committee of the University of Baghdad- College Of Science for women. The study was conducted in accordance with the guidelines of the ministry of Iraqi health and to the Helsinki Declaration. All patients had given a written informed consent before starting work.

Samples collection and processing:

Around 10ml of blood was collected using standard techniques from each participant in the fasting state. Blood samples were collected in a tube without anticoagulants at room temperature. Then serum was separated. Levels of elements in serum samples were measured through atomic absorption method, serum sample was taken into breakdown container and break down with 2ml of nitric acid and perchloric acid combination (with a ratio of 4:1) to eliminate interference from the organic matrix¹². Then the samples were incubated at least for 65 min with 120°C. The mixture was diluted with deionized Distilled water to brought the mixture volume to 10ml. Finally, the level of each element at the related wavelength was concluded via an atomic absorption device (novAA 350) in accordance with the guidelines¹⁰.

Quantification of the elements:

Elements in the samples (Fe, Zn, Cu, Se, Al, Si and Cr) were specified via novAA 350 flame atomic absorption spectrophotometer. All the measurements were performed in triplicate.

Statistical analysis:

Statistical analysis program (SPSS 25) have been performed for analysis the results. General descriptive statistic was used to describe the main findings; Student t-test has been used to identify the significant differences. The correlation has been studied used Spearman correlation. Dendrogram have been used to identify the similarity between variables Also, the receiver operating characteristic curve (ROC) analysis has been done to identify the cut off value for the parameters.

RESULT AND DISCUSSION:

The Contributors demographic factors:

The demographic characteristic associated to the brucellar spondylitis patients and matching healthy donor are summarized in Table 1.

Table 1 demographic characteristic for brucellar spondylitis patients and healthy donor.

Characteristics	Patient’s	Control
Number	41	21
Age
Range	20-60	20-60
Mean	41	43
Gender
Female	25	11
Male	16	10
Clinical examination
Fever	40	0
Back Pain	39	9
Fatigue	38	6
Sweating	33	0
Laboratory tests
Rose Bengal plate tests	30	2
IgG test	37	0
IgM test	33	0
X-rays	30	0
MRI examination	38	0

All voluntary subjects age were divided in to 4 categories according to the range 20-30, 30-40,40-50, and 50-60. A 56.1 % of patents with brucellar spondylitis were lied under the age group of 40-50, and 34.1 % were lied under age range 30-40. This distribution suggested that the infection diseases were not correlate with age. Chart 1 describe the age range distribution in to for categorize.

Chart 1: Depicts the age group of A: Control and B: brucellar spondylitis

Distribution of trace/toxic elements in serum samples:

Levels of trace/toxic elements in serum samples of control group and patient group were presented in table 2. Average concentration revealed dominant contribution for Fe (6ppm), followed by Cu (9.5ppm), Zn (9.3ppm), Se (151.9ppb), Al (12.7ppb), Si (5.3ppb) and, Cr (5.5 ppb) in brucellar spondylitis. All presented elements approved highly significant difference at 0.01 levels. Fe was significantly elevated in brucellar spondylitis followed by Cu, Zn, Se, Al and, Cr, whereas Si showed significant decrements in patients with brucellar spondylitis when compared to control subjects. The significant value that have been appeared in all the selected elements through statistical analysis using two - tailed student T Test, were considered as an interested result, as long as the diagnose of brucellar spondylitis still a big issue.

In general, some authors reported that brucellosis made alteration in level of copper, magnesium, zinc and manganese.¹⁰ The alteration in elements level in serum sample, appeared as a result of infection disease such as brucellosis via alteration or impaired liver metabolism.

Table 2: Statistical distribution parameters of selected elements levels in the serum (Fe, Cu and Zn in ppm unit) and, (Se, Al, Si, and Cr in ppb unit) of the brucellar spondylitis patients, and controls.

Parameter	Control	brucellar spondylitis
Parameter	Mean ± SD	Mean ± SD
Fe	1.9±0.16	6.0±0.16^**
Cu	0.18±0.015	9.5±2^**
Zn	1.19±0.04	9.3±1.7^**
Se	118.5±5.4	151.9±3.0^**
Al	7.85±0.39	12.7±0.54^**
Si	109.9±9.9	5.3±0.25^**
Cr	0.338±0.02	5.59±1.1^**

*level 0.05 (2-tailed), significant

**level 0.01 (2-tailed), significant

Elements showed vital role and prognostic significance in complex disorders leading to schizophrenia²², other study showed alteration in trace elements in diabetes patients²⁴

Correlation study.

The association between the parameters were studded through applying Spearman correlation, which is helped to identify the strong of correlation and the significant, table 3 shows the spearman correlation coefficient for the selected elements in the serum samples, All the elements were correlated positively; however, only Si is correlated negatively. Zn, Cu, and Cr showed very strong correlation.

Table 3: Spearman correlation coefficient of studied elements in the serum of all subjects. When r upper than 0.3 or lower than -0.3 the result is significant at p < 0.001.

Parameters	Fe	Cu	Zn	Se	Al	Si	Cr
Fe	1	.362**	.370**	.605**	.508**	-.840**	.366**
Cu		1	1.000**	.126	.352**	-.342**	1.000**
Zn			1	.136	.358**	-.352**	1.000**
Se				1	.152	-.674**	.132
Al					1	-.571**	.356**
Si						1	-.348**
Cr

* level 0.05 (2-tailed), significant

**level 0.01 (2-tailed), significant

The analysis of Multivariate

The corresponding CA (that describe the similarity via studied variables) based on Wardʼs method of studied variables was described in (chart 2A) for serum f control and (chart 2B) for serum of brucellar spondylitis. the diagram shown great bands of Cu-Cr, Cr- Fe, Fe- Zn, Zn-Al, on the other hand Se-Si revealed another strong cluster using data of control, Whereas, data of brucellar spondylitis showed very strong clusters of Cu-Zn, Zn-Cr, Cr-Fe, Fe-Si, Si-Al. The involvement of the most elements with similar cluster suggested that, the environmental contamination as well as to internal body metabolism plays a role in regulation of studied elements. This multivariate process encouraged changed elemental circulation and distribution design in the serum of brucellar spondylitis with the controls. Multivariate statistical analysis displayed noticeably different association of the elements in the brucellar spondylitis and control group. The study designated deep modification of the elemental points in the brucellar spondylitis; it might be disorders including brucellar spondylitis.

Chart 2 Cluster diagram of studied elements in the serum of A: control and B: brucellar spondylitis

Trace elements play some significant roles in human body, first the body required a smallest quantity of elements to be healthy, second it plays an important role in the immune system. Furthermore, some elements act as cofactors in many biological reactions. It is well known that the body need low concentration of these elements and, any alteration in their levels leads to disrupt several pathways in biological system. Make the body defenseless against pathogens²⁵. Some elements have a significant role in resistance to infectious diseases like Cu and Zn.²⁶. Current study showed that Fe, Cu, Se and Zn in patients with brucellar spondylitis had significant increment compared to that in the control group, Other studies were agree with this study except that Zn appeared decrease in its level compared with control group^27,28. In 2014 other researchers concluded that there were no alteration in the serum level of Cu and Zn in the patients with brucellosis when was compared to control group²⁹. Extra researches are important to identify whether the studied elements have an effect on disease severity and outcome. Assaying these elements level may be proposed as a complementary screening for Infectious diseases especially brucellar spondylitis. Infection disease such as brucellar spondylitis can affect of serum elements through altered of liver metabolism, one of the most common altered is synthesis of ceruloplasmin in the liver which affects copper level²⁸. The next interesting micronutrients is Cr, it has several roles in the body and, it cannot have synthesized inside the body, so it must have obtained from external source like a diet. The level of Cr showed increment in it is concentration compared to control; the alteration in Cr level may be as a results of involvement of liver in the systematic infection. There were no enough results in review regarding of the Cr concentration. Also, there are no studies in review regarding the involvement of the micronutrient Al, and Si in cases of brucellar spondylitis. In the current study Al showed significant elevation as compared to control group, whereas Si showed significant decrement, when was compared to control group.

Receiver operating characteristic analysis (ROC):

ROC has been applied to identified the pure cut off value for the tested elements using presented data, this analysis was produced via scheming the true positive which is recognized as sensitivity contrary to the false positive which is identified as (1- specificity). The relationship reveals the trade off between sensitivity and specificity of selected micronutrient Fe,Cu,Zn, Se, Al, Si and Cr this relationship translated to differentiate between brucellar spondylitis and control group, The beset cut off value were (2, 0.17, 1.3, 121, 7.5, 118, and 0.35) for Fe, Cu, Zn, Se, Al, Si and Cr respectively chart 3 showed the ROC curve for all the selected elements.

Chart 3 Receiver operating characteristic (ROC) for the selected elements (a: Fe, b: Cu, c: Zn, d:Se, e:Si, f: Cr and, g: Al)

CONCLUSION:

In the current investigation, we watched changed degrees of the essential and toxic elements and accepted the determination of components inside the serum can be helpful for assessing the brucellar spondylitis. Such alteration may impact be valuable signs to screen for and possibly display the decline of disease. The investigation of elements in serum of brucellar spondylitis and connected control subjects gives information to Orthopedic specialists about the significance of the change in the digestion of the elements.

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Received on 19.03.2021 Modified on 25.07.2021

Research J. Pharm. and Tech. 2022; 15(5):1933-1937.

DOI: 10.52711/0974-360X.2022.00321