Journal of Advanced Zoology
ISSN
:
0253-7214
Volume
44
Issue
S-5
Year 2023
Page
385
:
391
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385
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Chlamydiosis in Sheep: Immunological Examination And
Pathomorphological Changes
Navruzov N.I.
1*
, Aktamov U.B.
2
, Sayfidinov B.F.
3
1,2,3
Samarkand State University of Veterinary Medicine, Animal Husbandry and Biotechnology
*Corresponding author’s: Navruzov N.I.
Article History
Received: 06 June 2023
Revised: 05 Sept 2023
Accepted: 12 Oct 2023
CC License
CC-BY-NC-SA 4.0
Abstract
The article provides information on the stability of the immune system and path
morphological changes in chlamydia when using the formalin GOA vaccine
against chlamydia in sheep.
Keywords: Chlamydia, Immunoglobulin, Vaccine, Immunity, Immunophone,
Antigen, Microorganism, Receptor, Serotype
1.
Introduction
Relevance of the research
Chlamydiosis in small and big horned animals is well documented to inflict significant economic harm
to all farms in our nation. Investigations have shown that chlamydia is responsible for an average of 12
percent of abortions in animals. Up to 50% of miscarriages in farm animals are caused by chlamydia,
claim I.I. Nosov and A.A. Volkova. The care of sick animals and the efforts to stop the sickness cost a
lot of money.
According to the literature, the prevalence of chlamydia is 18-43.6 in the United States, 9-21.6 in
Canada, 16-24.8 in the Netherlands, 18-57 in France, and 14-46.4 in England. In Australia and Israel,
the prevalences are 6-26.2% and 3-34%, respectively.
Inflammation of the placenta, particularly the cotyledons, abortion in the second half of the cervix, the
birth of weak lambs and calves (often young animals), and inflammation of the lungs (pneumonia) are
all symptoms of the spreading, enzootic infection known as chlamydia.
Chlamydia abortus ovis, a member of the Chlamydiacea family and Chlamydiacea psittaci genus, is the
responsible party. Chlamydia is a parasite that may grow up to 250–300 nm in size. They are bacteria
with thick cell walls that carry DNA and RNA.
The chlamydia-causing agent has a complicated antigen structure with three antigenic centers that are
unique to the genus, species, and serogroup. Its gender is lipopolysaccharide because, like gram-
negative bacteria, it has a thermostable cell wall. The binding epitope, which establishes the specificity
of the genus, comprises of a unique receptor found in carbohydrates and an oligosaccharide molecule
made up of three monomers, which expresses antigenicity (G.A. Dmitriev et al., 1999). The particular
location of cysteine-rich amino acids in the protein membrane of species-specific determinants varies
between antigen serotypes.
The purpose of the research.
Stability of the effect of the GOA formalin vaccine against sheep
chlamydiosis on the immune system with IgM and IgG test kits developed by JV "UNIGEN" is the
main criterion of our experience to determine the effectiveness of the vaccine.
2.
Materials And Methods
Research was done at the VITI Regional Diagnostics, Microbiology, and Pathomorphology laboratories
as well as in the livestock complex in the Dehkanabad District of the Kashkadarya Region called
"Karakolchilik Shirkati named after M. Ibragimov."
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The quantity of immunoglobulins and how they affected the infections affected the div's ability to
fight against germs. Farm animals almost never have immunoglobulin-E or immunoglobulin-D
identified (F.J. Bourne et al. 1978). The first step of immunological responses is marked by the
appearance of IgM from macroglobulins. The primary immunoglobulin in blood serum is IgG, which
comes in two varieties: IgG
1
and IgG
2
. The primary cellular components of the div, in addition to
immunoglobulins, are macrophages (monocytes), active T and V lymphocytes, and immunoglobulins,
which enable the div's resistance against viruses and microbes (Green S.A., Albulov A.I., Ruban E.A.,
Green A.V. ). The morphological and pathological status of the div's tissues and cells is negatively
impacted by the antibiotics used to treat the disease. It is important to remember that polyclonal
activation syndromes are what lead to false positive outcomes in such enzymatic and sequential
processes. The creation of specific defensive protein enzymes against diverse foreign antigens entering
the div is simultaneously stimulated by superantigens, which are present in the individual
(ontogonistic) phase of the animal div, and V cells respond in a particular manner. In actuality, these
activities manifest as a simultaneous, non-specific rise in the titer of antigens to several infections.
According to literature sources, technical flaws in the production of the response as well as
immunodeficiency circumstances may be to blame for false negative findings in the detection of
antigens. A specific apparatus (colorimeter) was used to measure the color intensity of the liquid in the
tablet wells after the reaction had halted, and a special gadget was utilized to compute the results. The
control samples' optical densities were compared, and the analysis's findings underwent mathematical
processing. It was determined that the sample contained more particular chlamydial antibodies the
greater the optical density in this well.
Antigen was preadsorbed on the well walls of 96-well polystyrene tablets for IEA. The well of the tablet
was filled with the blood serum that would be evaluated. In this instance, the homologous antibodies
join after attaching to the previously adsorbed antigen. During washing, chlamydia antibodies that are
not bound are eliminated. The well was subsequently filled with enzyme-labeled antibodies (chlamydia
antibodies) directed to rabbit or other animal immunoglobulins. If any chlamydial antibodies are found
in the blood serum under examination, they act as antigens at this point and mix with chlamydial
antibodies found for the enzyme. After washing, a chromogenic (coloring) material was applied,
allowing for the possibility of accounting for the reaction on the growth of staining in the wells. Since
the number of chlamydia antibodies and the amount of enzyme are quantitatively identical, the intensity
of staining is proportional to both. The optical density of the liquid in the wells was measured and
compared to the optical density of the control sample to determine the antidiv concentration per unit
volume. Results were calculated using optical density units. For taking IEA findings, levels of normal
indicators, and pathological indicators into consideration, each test system has its own indicators.
Results of immunoenzyme analysis are based on them.
IEA was conducted using "Socorex" dispensers, ELx405 microplate cleaning apparatus, and ELx808
microplate automated analyzers. Using a computer and the Bio-Tek KC4TM software, the reaction's
findings were electronically (and hence electronically) interpreted.
In an experiment, the serological and immunological responses in sheep that had received the
vaccination were examined to determine the efficacy of the IgM and IgG test kits created by "UNIGEN"
and "XEMA" LLC as preventative measures.
Ten sheep were administered two subcutaneous injections of the "emulsified vaccine against
chlamydiosis" in experimental group I.
Only one dose of the "Chlamydiosis vaccine" was administered to the II comparable group of 10 heads.
Group III (10 heads) served as the control group, receiving no medication. Based on the farm
veterinarian's anamnesis data and taking into consideration the fact that the lambs that miscarried and
gave birth the previous year were not viable, the sheep chosen for the study were decided.
3. Results and Discussion
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We studied immunoenzymatic analysis (IEA) based on the short-term detection of the reaction against
the provoking antigen or the specific antidiv formed against it. Although the IEA method allows
differentiating between infected and vaccinated animals in the case of small horned animals vaccinated
against chlamydia in the experiment, we used the serological (KBR) method in order to determine that
it is easier, faster, and more convenient to diagnose compared to the immunological method (Table 1).
Table 1
Optical Density Values
1
2
3
4
5
6
7
8
9
10
11
12
A
1,413
0,062
0,867
0,245
000
000
000
000
000
000
000
000
B
1,451
0,069
0,962
000
000
000
000
000
000
000
000
000
C
0,257
0,136
0,164
000
000
000
000
000
000
000
000
000
D
0,224
0,228
0,135
000
000
000
000
000
000
000
000
000
E
0,096
0,923
1,899
000
000
000
000
000
000
000
000
000
F
0,092
1,926
0,892
000
000
000
000
000
000
000
000
000
G
1,517
0,038
0,055
000
000
000
000
000
000
000
000
000
H
1,578
0,052
0,034
000
000
000
000
000
000
000
000
000
The samples in the first pair (A1 and V1) are the optical density standards for negative samples, while
the following pair (S1 and D1) are for positive blood samples, as can be seen from the table's data.
Blood samples from cells G1, H1, F2, and E3 were positive, whereas blood samples from cells E2, A3,
and B3 were doubtful, according to the examination of 21 analyzed blood samples.
Based on the high specificity and sensitivity of "antigen-antidiv" immunological responses, enzyme
immunoassay is a laboratory experiment. IEA is made up of two distinct parts: immunological and
enzymatic processes. The immunological response (bacteria and viral molecules) acted as an antigen
and a binding site for antibodies.
Additionally, the outcomes of the immunological reaction may be observed and quantified thanks to
the enzymatic reaction. In order to manage the epizootological situation and ascertain the general
immunophone in the farm of "Karakolchilik shirkat named after M. Ibragimov" LLC, Kashkadarya
area, Dehqonabad district, immunoenzymatic analysis (IEA, ELISA) reaction was utilized as an
immunobiological approach. Prior to starting the reaction, precautions were taken to assure compliance
with the rules and guidelines for biological safety in the laboratory.
Immunoenzymatic analysis (IEA or ELISA) for detection of IgG-specific antibodies against chlamydia
in the blood sera of cattle and small horned animals. "Chlamydia IgG-IFA" IgG antibodies to the
causative agent of chlamydia "Chlamydia IgG-IFA" developed in cooperation with the joint ventures
"UNIGEN" and "XEMA" LLC. "Set of reagents for detection with" ("Nabor reagentov dlya
immunofermentnogo vyyavleniya IgG antitel k vozbuditelyu chlamydiosis krupnogo i melkogo
rogatogo skota") was performed using the test system (Figures 1, 2). and was performed following the
general rules for performing the IEA reaction.
Figure 1. IgM-immunoglobulin
Figure 2. IgG-immunoglobulin
Table 2
Immunological analysis of chlamydiosis vaccine.
Types of analysis
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Groups
Number of
animals
C-reactive protein
(normally 0.1-0.3 mg/l)
IgM (normally 0.4-
2.3 mg/l)
IgG (normally 7-
16 mg/l)
Experimental
group I
10
0,32±0,025
2,8±0,23
17,1±0,96
Experimental
group II
10
0,287±0,015
2,04±0,143
16±1,144
Experimental
group III
10
0,108±0,0058
0,286±0,022
7,3±0,46
In experimental group I, it was discovered that the level of C-reactive protein was 1.55 times greater
than the average. It was discovered to be 1.13 times higher than the first comparison experimental group
and at the standard level in the II comparative experiment group. It was discovered that experimental
group I had a result that was 1.69 times higher than the norm and group II had a result that was 1.21
times higher when determining the course of the disease in a chronic condition according to the change
of IgG. IgM and IgG levels were found to be particularly high in the first group, with just a slight
difference from the II group and a significantly greater effect from the III group.
"M.Ibragimov" firm farm, Dehkanabad district, Kashkadarya area; and "Jizzakhlik" m.f.y., Jizzakh
region. By using the enzyme-linked immunosorbent assay technique, IgM markers were found in 11
samples of citizen blood as well as aborted babies that were 4 months old. In these farms, the
parenchymatous organs, caruncles, and cotyledons around the uterus of two sheep were histologically
analyzed.
In collaboration with the staff of the local diagnostics and microbiology laboratories, pathological
samples collected from farms were thoroughly examined pathologoanatomically in order to research
the epizootic status of chlamydia illness in the areas of our country. The following pathological
disorders were primarily seen in these animals when they were being examined.
During pathologoanatomical investigations, atelectasis, air buildup, and blood stagnation were seen in
the lambs' lungs. It was discovered that the heart muscles were relaxed, the auricles were developing
dotted and spotted hemorrhages, and the volume of blood in the heart chambers was extremely low.
These alterations suggest heart failure, and other internal organs have also been impacted by this
process. The liver's consistency has hardened, and various-sized abscesses and necrosis foci have
appeared. The kidneys' surface has point hemorrhages, the spleen is loose, and there are swellings in
certain places on its surface. There is a buildup of gas and catarrhal irritation, and the undigested milk
in the rennet has transformed into a floating mass.
All of these animals' pathological samples were fixed after which they were embedded in paraffin, cut
into histosections, and stained with hematoxylin-eosin. The following findings were found when the
produced histological slices were viewed under a microscope.
Hyperemia of the interalveolar barriers, enlarged pulmonary artery walls, and polymorphic cellular
infiltrates are also seen. As a result of these modifications, different pneumonias, or inflammations with
tiny foci, foci, and inflammations related to foci, developed (Fig. 8).
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Figure 8. Hyperemia of the interalveolar barriers in the lungs and various cellular infiltration.
On the basis of responses of arteries and cells, the stroma, the connective tissue that forms the
foundation of the heart, swelled. Different regions of the myocardium also developed collagen and
elastic fibers, and the cardiomyocytes clearly underwent dystrophy (Fig. 9).
Figure 10. Hydropic dystrophy of hepatocytes.
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Figure 9. Heart stroma swelling and cardiomycetic dystrophy.
Due to the change in the permeability of the vessel walls, hepatocytes of the liver develop various
degrees of hyaline droplet and watery (hydrophic) dystrophies. There are also inflammatory reactions
in the tissues of the liver tract (Fig. 10).
An infectious inflammatory process is present in the liver if lympho-macrophage infiltrates with a few
plasma cells start to develop.
As a result, tissue degradation, hemodynamic abnormalities, alterative processes, immunopathological
processes, and systemic inflammation were all symptoms of small horned animal chlamydiosis.
4. Conclusion
It From February to May, Samarkand region animal farms tested positive for 8.2 percent chlamydia;
Kashkadarya region farms tested positive for 6.9 percent chlamydia. Despite the excellent sensitivity
and accuracy of the responses in both cases when diagnosing chlamydiosis by serological (KBR) and
immunological (IFT) approaches, it was discovered that immunoenzymatic analysis was simple to
perform. The chlamydiosis-causing agent's cultural, morphological, tinctorial, virulence, biochemical,
and pathogenic properties were found in several cattle farms with a focus on breeding in the
Kashkadarya and Samarkand areas. It was shown that oleandomycin is sensitive to doxilox, teliosin,
oxacillin, and gentamicin are not sensitive, and erythromycin is less sensitive than these antibiotics in
treating chlamydia.
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