[Gazonof]

Сообщение от АНД

а самі на всякий случай не приймаєм антибіотики, а вдрух сьогодні грип підчепим, а ми не вживали таблетку,якось так

А як же щеплення наприклад

По теме, выводы делайте сами.
Как под спойлер сунуть?
Или читайте сами оригинал file:///C:/Users/scilla/Downloads/84-114-PB.pdf
страница 61

Науковий вісник ЛНУВМБ імені С.З. Ґжицького, 2018, т 20, № 83
Scientific Messenger LNUVMB, 2018, vol. 20, no 83
61
Науковий вісник Львівського національного університету
ветеринарної медицини та біотехнологій імені С.З. Ґжицького
Scientific Messenger of Lviv National University
of Veterinary Medicine and Biotechnologies
ISSN 2518–7554 print doi: 10.15421/nvlvet8312
ISSN 2518–1327 online http://nvlvet.com.ua/
UDC 619:638.15-08
Quantitative changes in hemogram of bees using probiotic «Enteronormin»
S.F. Tushak
Zhytomyr National Agroecological University, Zhytomyr, Ukraine
Article info
Received 06.01.2018
Received in revised form
19.02.2018
Accepted 27.02.2018
Zhytomyr National Agroecological
University, Korolova Str., 39,
Zhytomyr, 10025, Ukraine.
Tel.: +38-063-178-11-13.
E-mail: svetlana.tushak@ukr.net
Tushak, S.F. (2018). Quantitative changes in hemogram of bees using probiotic «Enteronormin».
Scientific Messenger of Lviv National University of Veterinary Medicine and Biotechnologies.
20(83), 61–65. doi: 10.15421/nvlvet8312
The article considers the main quantitative changes in hemolymph cells under the action of the drug
«Enteronormin» when fed with 50% sugar syrup during 10 days compared with the changes in the control
group that received only sugar syrup. On the 3rd day, the hemogram of the bees that received the drug did
not differ from the bees that received the sugar syrup only. Positive dynamics in the quantitative composition of the hemolymph was observed on the days 7–10. The number of phagocytic cells was 1.38% less than
in the control group, and 2.0% of neutrophils less and 0.63–1.13% of eosinophils. The number of secretory
cells increased, in particular, gliwocytes by 1.17% and 1.62%, respectively. It is noted that the medication
«Enteronormin» has been extended to the extent of 3 days not a positive effect on the immune system of
bees. Therefore, the drug «Enteronormin» should be fed for at least 7 days, taking into account the quantitative changes in hemocytes. On the twenty-first day the effect of the drug was preserved. The most appropriate term for drug feeding with the aim of activating the cellular immunity is defined to be 7–10 days.
Key words: bees, probiotik, «Enteronormin», hemogram, hemocytes, hemolymph, eosinophils,
progemocytes, spherulocytes, enocitoid.
Introduction
The state of the immune system is important for a living organism homeostasis formation. Immune status determines the effectiveness and consistency of all systems
and elements of immunity (Luss et al., 2000; Genersh and
Aubert, 2010). The immunity of honey bees ensures the
resistance of the bee family to the pathogenic microorganisms and their waste products. The formation of immunity
involves the entire organism of bees as an integral system
in which all the protective mechanisms are interconnected
(Artemenko and Skrypnik, 1984; Boman and Hultmark,
1987; Genersh and Aubert, 2010). The internal protective
mechanisms include the form elements of hemolymph
and fatty body, as well as the antimicrobial properties of
plasma, fluids and tissues of the insects’ body. The protective properties of form elements’ cells are called cellular immunity, while the protective properties of liquids –
humoral immunity (Boman and Hultmark, 1987; Glinski
and Jarosz, 2000). In the normal cell the composition of
hemolymph varies depending on the breed, season of the
year, conditions of keeping. In general, numerous studies
on the factors of immunity in bees have shown that the
stimulation of their activities causes complex biochemical
changes in their organism, which, according to their specificity and efficiency are less significant than the analogous changes in the organisms of warm-blooded animals.
Thus, any stage of bee development, its age and physiological state are characterized by a specific hemogram.
Hemograms are very sensitive to physiological changes in
the organism of insects. The hemogram of bees’ hemolymph is used for early diagnosis of diseases, as well as
for determining the effectiveness of the use of a particular
drug (Artemenko and Skrypnik, 1984; Boman and Hultmark, 1987; Glinski and Jarosz, 2000; Sapcaliu et al.,
2010).
The use of stimulants to prevent infectious diseases
requires a more thorough examination of their effectiveness. According to the theory stating that all hemocytes
have one source of origin (progemocytes) and claiming
the ability of different cell types to transform into amoeba-like state for phagocytosis, cytological changes due to
the immune drugs feeding can be analyzed. Moreover it is
important to mention the study by V.P. Tishchenko
(1986) proving that the insect hemogram reflects changes
in their organism according to the physiological norm; in
terms of hemocytes it is possible to evaluate their «fattening», to carry out an early diagnosis of diseases, parasite
infections and cases of insect intoxication (Tyshhenko,
1986; Glinski and Jarosz, 2000; Sapcaliu et al., 2010).
Науковий вісник ЛНУВМБ імені С.З. Ґжицького, 2018, т 20, № 83
Scientific Messenger LNUVMB, 2018, vol. 20, no 83
62
One of the factors that significantly influences the
change in cellular composition is the use of various biologically active drugs, in particular, probiotics. The beekeepers of Ukraine began to use the drug «Enteronormin», a probiotic, or a symbiotic, containing viable lactic
acid and sporogenic bacteria in a unique symbiosis (Enterococcusspp., Lactobacillus spp., Bacillus tubilissp.)
with the addition of prebiotic chitosan and peptone. Lactic
acid bacteria in the drug Enteronormin are specially selected to have the maximum ability to extract lactic acid,
useful enzymes, essential vitamins, stimulators and modulators of the immune system, and other biologically active
substances.
Enteronormin actively forms normal microflora, participates in activating the protective forces of the bee's
organism, stimulates the development and productivity of
bee colonies, exhibits antagonistic (suppressing) activity
against a wide variety of pathogenic bacteria and fungi
(Rekomendacii po primeneniju …).
Analysis of recent research. Veterinary doctor Rubel
I.S. from the company TD «Geotek» carried out the study
on the life expectancy of bees using the probiotic
«Emprobio». The effect of the probiotic on the hemolymph composition was not investigated. Domanska T.
studied the effect of probiotics Vetom-1.1 and Vetom-3
for the treatment of bees families suffering from ascosferosis (Rubel' et al., 2013).
The objective of the research. To study the effect of
probiotic «Enteronormin» on the hemolymph of bees. To
identify the positive effects of the drug on the hemogram
of bees, depending on the duration of treatment.
Materials and methods
The main tasks were: 1) to define the optimal term of
the drug Enteronormin feeding taking into account the
changes in hemolymph; 2) to detect in laboratory conditions the major changes that may occur in the composition
of hemolymph when feeding the probiotic drug Enteronormin, depending on the term of the drug feeding; 3) to
identify presence of the drug prolonged action.
The analysis of the effect of the drug «Enteronormin»
on the hemolymph of bees was carried out in the laboratory of the Department of Microbiology, Pharmacology and
Epizootology of the Zhytomyr National Agroecological
University during April–June 2017. The bees were kept in
a thermostat with a passive ventilation system, where the
temperature of 26 °С and a relative humidity of 35–70%
were maintained. 100 species treated by the drug in a ratio
of 1:5 with 50% sugar syrup were kept in entomophylic
containers. Prior to the treatment, for the activation of
microorganisms the required amount of the drug was
dissolved in an aqueous solution of «Jodis + Se» in a
proportion of 1 to 5 and infused during 16 h. The drug
was prescribed orally during 3–10 days.
The hemolymph analyses were carried out on the 1st,
3rd, 7th, 10th and 21st with the modified method «SNAU,
Vet-NSC Prokopovicha» in order to detect the effect of
the drug on hemolymph of bees. This effect will indicate
the activation of immune processes in the body of bees.
The data was processed using the statistics method and
Excel.
Results and discussions
The microscope analysis of smears of individual samples of bees hemolymph, showed that at the beginning of
the research (before using the drug), the quantitative indices of hemocytes constituted: proleucocytes 21.69%,
neutrophil phagocytes 28.56%, eosinophilic phagocytes
23.56%, spherulocytes 23.38%, enocitoid 2.75%.
The results of treatment with «Enteronormin» are presented in Table 1. Analyzing the data from Table 1, it
should be noted that the number of hemocytes in bees is
unstable and varies with the time depending on the environmental factors.
Table 1
Quantitative changes in bees hemolymph indices
Groups of
bees Proleucocytes Neutrophilic phagocytes Eosinophilic phagocytes
Before the
experoment 21.69 ± 2.27 28.56 ± 2.78 23.56 ± 2.99
On control drug сontrol drug control drug
3-d day 19.20 ± 1.75 19.0 ± 2.27 29.10 ± 1.37 29.25 ± 0.76 24.30 ± 1.83 24.75 ± 2.71
7-th day 18.25 ± 1.67 18.75 ± 1.28 29.63 ± 1.68 28.25 ± 1.39 24.38 ± 2.26 23.75 ± 2.43
10-th day 18.0 ± 0.75 19.13 ± 0.99 30.0 ± 1.07 28.0 ± 0.76 24.88 ± 0.64 23.75 ± 1.04
21-st day 17.5 ± 1.07 18.38 ± 0.92 31.0 ± 0.93 29.13 ± 2.85 25.13 ± 1.13 24.13 ± 0.64
Groups of
bees Enocitoids Spherulocytes
Before the
experiment 2.75 ± 1.77 23.38 ± 2.82
On control drug control drug
3-d day 2.5 ± 1.78 2.37 ± 1.68 25.3 ± 1.66 24.87 ± 2.23
7-th day 2.75 ± 0.83 3.4 ± 1.68 24.63 ± 1.92 25.8 ± 1.64
10-th day 2.75 ± 0.71 3.13 ± 0.83 24.375 ± 0.52 26.0 ± 0.76
21-st day 2.13 ± 0.83 2.88 ± 1.13 24.25 ± 1.67 25.5 ± 1.19
Науковий вісник ЛНУВМБ імені С.З. Ґжицького, 2018, т 20, № 83
Scientific Messenger LNUVMB, 2018, vol. 20, no 83
63
When comparing the hemolymph index, analysed on
the 1st day before the beginning of the experiment with
control group on the 3rd, 7th, 10th and 21st day of the
experiment, an increase in phagocyte cells was observed –
neutrophil by 0.54%, 1.06%, 1.44% and 2.44% respectively, eosinophils by 0.94%, 0.82%, 1.32% and 1.57%,
indicating the activation of hemocytes phagocytic
reactions and the reduction of new cells reproduction,
probably due to the unnatural laboratory conditions and
aging. In this case, the comparison of cellular hemolymph
response during and without the use of stimulants demonstrates the emergence of adaptive mechanisms in
experimental groups kept in unnatural laboratory
conditions (Fig. 1). Taking into account the conditions of
bees maintaining in the thermostat (approximating the
natural conditions) a rapid increase in indicators is a
norm.
The comparison of the microscopy results of the hemolymph smears of control group and the bees receiving
by the drug is presented in Figure 1 and Figure 2. It
should be indicated that on the 3-d day the positive
changes in the hemogram were not observed, in particular, the quantity of proleucocytes remained constant. The
number of phagocytes increased by 0.15%, neutrophilic
phagocytes and osinophilic phagocytes by 0.25%. Moreover, the decrease in secretory cells encitoids and spherulocytes was noted by 0.13% and 1.57% respectively. The
results mentioned above indicate that the use of the drug
during 3 days does not allow implementing the active
stimulation of bees’ organism.
Figure 1. The dynamics of quantitative changes of phagocytes in the hemolymph of bees from control group
and those treated by «Enteronormin»
Figure 2. The dynamics of quantitative composition of hemolymph secretory cells of bees treated
by «Enteronormin» and control group.
It should also be noted that positive changes in the
hemogram were observed on the 7th and 10th days of the
experiment. Thus, compared with the control group, the
quantity of proleucocytes was 0.5% and 1.13% higher
respectively, indicating a positive dynamics of the immune processes in the organism of bees, in particular the
ability of the body to produce «fresh» cells. The number
of phagocytic cells was decreased in comparison with the
control, therefore it can be concluded that the immune
system of bees does not perceive the drug as an alien
Науковий вісник ЛНУВМБ імені С.З. Ґжицького, 2018, т 20, № 83
Scientific Messenger LNUVMB, 2018, vol. 20, no 83
64
body, but rather as a stimulant. The number of spherulocytes increased by almost 2% compared with control,
moreover there appeared young forms of spherulocytes
with small vacuoles.
Taking into account the results of the research, it
should be noted that the feeding of the drug within 7–
10 days gives positive results. From the 10th till the 21st
day only sugar syrup was fed to bees. At the end of the
experiment (the 21st day), smears of hemolymph of bees,
constantly receiving sugar syrup and those receiving the
drug up to the 10th day, were examined. It can be stated
that the quantitative indices of hemolymph cells on the
21st day taken from the bees fed only by sugar syrup
indicate the aging of the bees, so the proleucocytes constituted 17.5%, phagocytes – neutrophils 31% and eosinophils 25.13%, spherulocytes – 24.25%. The reason for the
faster aging of the bees’ organism is the conditions of
keeping, since they are as close as possible to the natural,
but not equal to them.
Regarding the morphologic changes in hemolymph
cells of bees, we observed significant changes in hemolymph of bees in the control group. Clearly defined mature neutrophil phagocytes, manifested by the formation
of pseudopodia at the opposite poles of the cell and their
extension, cells acquired spindle-like shape, the nucleus
shifted from the center of the cell to the periphery. These
cells are highly specialized, actively carrying out phagocytosis and encapsulation of foreign substances. These
cells were observed on days 7–10 of the experiment in
bees receiving a sugar syrup only. The results were different for the bees receiving the drug (Fig. 3).
Fig. 3. Mature forms of phagocytes

In the hemolymph of bees receiving the syrup the
mixed with the drug, vivid morphological changes in the
spherulocytes were observed. Thus, in hemolymph, the
young forms of cells characterized by clear contours and a
dense centered nucleus, the cytoplasm filled with small
vacuoles, in contrast to the old, poorly colored, losing
clarity of outlines, with the nucleus shifted to the periphery and often loose were detected (Fig. 4–5).
Figure 4. Young spherulocytes Figure 5. Old forms of spherulocytes
Quantitative indices of hemolymph of bees that were
fed the drug up to the 10th day were at the level of days
5–7, therefore we can claim that the drug «Enteroronorm»
stimulates the immune processes in the body of bees even
after completing the feeing of the drug – the aging of the
body is slower, the immune system is more stable to unfavorable environmental factors, indicating the prolonged
effect of the drug.
On the 21st day there appeared a tendency towards thr
stabilization of the quantity of proleukocytes (precursor
cells) in the experiment with Enteronormin, which
indicates a slow but prolonged effect of this drug on the
body of bees and their hemolymph.
Quantitative and qualitative changes in hemocytes of
spring bees generation during their stay in laboratory
conditions and the application of working doses of
enteronormicum, indicate the dynamism of hemocytic
indices of honeybee bees hemolymph and confirm the
importance of cytological analysis for rational choice of
immune drugs.
Conclusions
Feeding «Enteronormin» during 3 days did not show
an effective result in hemolymph rates, since no positive
changes in hemolymph were detected.
Effectiveness of the drug «Enteroronorm» is actively
manifested in feeding it with sugar syrup during 7–10
days, characterized by positive changes in hemolymph
and the activation of the cellular immunity of bees.
The effect of «Enteronormin» is preserved during 21
days, the immune system of bees becomes more resistant
to unfavourable environmental factors, the average life
expectancy of bees in containers increases by 2–3 days
compared with the control group. That confirms prolonged action of the drug and the activization of immune
processes in the organism of bees.
The perspective of future research. The next stage of
the reseach will be aimed at confirmation of the drug
«Enteronormin» efficiency if used in field conditions as
well as defining the degree of immunity activation using
the gemolymph analysis. Determination of the effect of
Науковий вісник ЛНУВМБ імені С.З. Ґжицького, 2018, т 20, № 83
Scientific Messenger LNUVMB, 2018, vol. 20, no 83
65
the drug on the hygienic behavior of bees. Improvement
of the method of probiotic «Enteroronorm» application
depending on seasons of the year and epizootological
situation at the place of use.
References
Artemenko, L.P., & Skrypnik, E.I. (1984). Immunitet u
pchel. Pchelovodstvo. 11, 18–19 (in Russian).
Boman, H.G., & Hultmark, D. (1987). Cell-free immunity
in insects. Annual Review of Microbiology. 41, 103–
109. doi: 10.1146/annurev.mi.41.100187.000535
Genersh, E., & Aubert, M. (2010). Emerging and reemerging viruses of the honey bee
(A. mellifera). Veterinary Research. 41, 54.
doi: 10.1051/vetres/2010027
Glinski, Z., & Jarosz, J. (2000). Problems of immunosuppression and immunotoxicology in respect to the honeybee protection against microbial and parasitic invaders. Apiacta. 35(2), 65–76.
Luss, L.V., Nekrasov, O.V., Puchkova, N.G., Bharvadzh,
A.V., & Bharvadzh, L.A (2000). Іmunologіja. 5, 34–
38.
Rekomendacii po primeneniju veterinarnogo
probioticheskogo preparata «Еnteronormin» v
pchelovodstve Rezhim dostupa:
http://aikam.com.ua/ru/goods/view/9002930/all/entero
normin--unikalnyy-probiotik-dlya-pchelovodstva/ (in
Russian).
Rubel', I.S., Perebejnis, A.V., & Rzhevskaja, V.S. (2013).
Vlijanie mikrobiologicheskogo preparata
«Jemprobio®» na uvelichenie prodolzhitel'nosti zhizni
rabochih pchel. Jekosistemy, ih optimizacija i ohrana.
9, 215–220. Rezhim dostupu:
http://nbuv.gov.ua/UJRN/ecooo_2013_9_22 (in Russian).
Sapcaliu, A., Pavel, C., Savu V., Căuia, E., Matei, M., &
Rădoi, I. (2010). Biochemical and Cytological Investigations on Haemolymph of Apis Mellifera Carpathica Bee
in Stressful Conditions. Bulletin UASVM Animal Science and Biotechnologies. 67(1–2), 313–320. doi:
10.15835/buasvmcn-asb:67:1-2:5317
Tyshhenko, V.P. (1986). Fiziologija nasekomyh. Ucheb.
posobie dlja studentov un-tov, obuchajushhihsja po
spec. «Biologija». M.: Vyssh. shk. (in Russian).