@phdthesis{
author = "Tarić, Elmin",
year = "2022",
abstract = "Doktroska disertacija, Štetoĉine i patogeni predstavljaju najĉešće uzroke gubitaka pĉelinjih društva.
Ameriĉka truleţ (AFB) je bolest pĉelinjeg legla, uzrokovana bakterijom Paenibacillus.
larvae, White (1906), moţe se smatrati glavnom pretnjom po zdravlje pĉela, s obzirom
da je reĉ o panzootskoj bolesti koja se vrlo brzo širi, ne samo iz košnice u košnicu, sa
pĉelinjaka na pĉelinjak iz regije u regiju, već i iz drţave u drţavu i šire. Evropska truleţ
je bakterijska bolest pĉelinjeg legla uzrokovana sa više bakterisjkih vrsta gde dominira
Melissococcus plutonius. Ova bolest je jako raširena i predstavlja veliki problem u
pĉelarstvu smanjujući proizvodne rezulte pĉelinjih zajednica. Kreĉno leglo (CHB) je
infekcija izazvana gljivicom Ascosphaera apis izazivajući gubitke komercijalno gajenih
pĉelinjih zajednica, naroĉito u kombinaciji sa vrstama mikrosporidija roda Nosema i
virusom mešinastog legla. Nozemoza je najĉešća bolest meĊu odraslim pĉelama
uzrokovana mirkosporidijama Nosema apis i N. ceranae i ĉesto dovodi do ekonomskih
gubitaka u pĉelarstvu. Virusne infekcije pĉela, koje su već odavno dostigle razmere
panzootije, ugroţavaju zdravlje pĉela i predstavljaju konaĉne egzekutore pĉelinjih
zajdnica. Pĉelinji krpelj Varroa destructor je glavni vektor skoro svih, a naroĉito
virusnih infekcija pĉela, koje su postale ozbiljan problem ne samo za komercijalno
gajene, već i za pĉelinje zajednice u divljini, zahvaljujući upravo krpelju V. destructor
kao fiziĉkom i biološkom vektoru Na osnovu svega navedenog jasno je da je
zdravstveno stanje društava u komercijalnom pĉelarstvu izloţeno velikim rizicima, a
njihovo ispitivanje i tretman predstavljaju veliki izazov kako za istraţivaĉe tako i za
pĉelare.
Bolesti odraslih pĉela i pĉelinjeg legla u komercijalnom pĉelarstvu najĉešće su
posledica energetskog stresa, nastalog zbog neadekvatne prihrane pĉelinjih zajednica,
prvenstveno prevelikom upotrebom sećernog sirupa. U osnovi energetskog stresa je
oksidativni stres pĉelinjih zajednica koji se moţe definisati kao disbalans izmeĊu
proizvodnje reaktivnih oblika kiseonika i antioksidativne odbrane. Reaktivni oblici
kiseonika (ROS) negativno utiĉu na ćelijske funkcije i stvaraju se tokom oksidoredukcionih metaboliĉkih procesa u ćeliji. Oni su ukljuĉeni u regulaciju razliĉitih
mehanizama, intercelularne signalizacije, a imaju i baktericidno dejstvo.
Medonosne pĉele, kao i ostale ţivotinje, razvile su niz enzimskih mehanizama
kojima se odupiru oksidativnom stresu, uklanjajući slobodne radikale. U ovim
procesima vaţnu ulogu umaju: superoksid dismutaza (SOD), enzim prisutan u citosolu i
mitohondrijama; katalaza (CAT) prisutna u peroksizomima glutation S-transferaza
(GST), glutation S transferaza (GST), peroksidaza i tireodoksin/tireoreduktazni sistem.
Imunitet pĉela, kao i imunitet u opšte, obuhvata kompleksan sistem koji ima cilj da
obezbedi oĉuvanje zdravlja kao i opstanak organizma na osnovu mnogih mehanizama
odbrane od patogena i drugih štetnih noksi (fiziĉkih i hemijskih). Medonosna pĉela svoj
socijalni imunitet ostvaruje uz pomoć bihejviorlanih mehanizama koji pomaţu zajdnici
u odbrani od patogena. Ulaganje u individualni imunitet ima visoku materijalnoenergetsku cenu za društvo, pa su udruţivanjem pĉela (razvoj socijlanog ustrojstva)
uspostavljeni zajedniĉki socijalni mehanizmi odbrane koji omogućavaju pĉelinjem
društvu efikasniju i ekonomiĉniju borbu protiv agenasa spoljašnje sredine. Za socijalni
imunitet pĉela od znaĉaja je enzim glukoza oksidazi (GOX). Ovaj enzim je produkt
pĉelinjih egzokrinih ţlezda, a uloga mu je u „konzervaciji― pĉelinjuh proizvoda, meda i
perge, tako spreĉavajući njihovo kvarenje. GOX uĉestvuje u procesu katalize -D
glukoze do glukonske kiseline i vodonik-peroksida. Vodonik-peroksid ima antiseptiĉka
svojstva, tako doprinoseći dodatnoj zaštiti i socijalnom imunitetu pĉela.
Cilj istraţivanja je bio da se utvrdi: da li izmeĊu tradicionalno i savremeno
gajenih pĉelinjih društava postoje razlike u pogledu prisustva patogena i pojave bolesti
pĉelinjeg legla, uz analizu bioloških (ekspresija GOX gena i faktora oksidativnog
stresa) i antropogenih faktora (menadţment u pĉelarstvu) na pojavu praćenih bolesti
pĉelinjeg legla i odraslih pĉela.
Terenski deo istraţivanja je obavljen na prostoru Pešterske visoravni (opština
Sjenica, Srbija) na 144 asimptomatske pĉelinje zajednice. Uzorci odraslih pĉela i
pĉelinjeg legla su uzeti iz savremenih (DB) košnica i tradicionalnih košnica „trmki―,
kako bi se utvrdila zastupljenost patogena legla (Paenibacilus larvae, Melissococus
plutonius, Ascosphera apis, virus mešinastog legla, tripanozoma Lotmaria passim i
Crithidia mellificae) i odraslih pĉela (virus akutne paralize pĉela, virus hroniĉne
paralize pĉela, virus deformisanih krila pĉela, tripanozoma Lotmaria passim i Crithidia
mellificae,mikrosporidija Nosema apis i Nosema ceranae ) kao i za utvrĊivanje nivoa
oksidativnog stresa analizom aktivnosti enzima: SOD, CAT, GST i koncentracije
malondialdehida (MDA). TakoĊe, iz istih košnica uzeti su uzorci za procenu ekspresije
GOX gena kao parametra socijlanog imuniteta.
Detekcija patogena je obavljena izolacijom bakterija kao i upotrebom PCR i
real-time PCR, u skladu sa standardima OIE. Rezultati istraţivanja su pokazali da je
meĊu komercijalno gajenim društvima P. larvae bio zastupljen u 16,67% uzoraka, A.
apis bio prisutan u 15,83% uzorka, dok je SBV detektovan u 96,67% uzoraka. MeĊutim
u leglu pĉela gajenih u tradicionalnim trmka košnicama naĊen je samo SBV u 33,33%
uzoraka. Dalja istraţivanja su utvrdila da su kod odraslih pĉela u komercijalno gajnim
društvima bili signifikantno (p<0,001) više zastupljeni virusi: ABPV, CBPV i DWV
(83,33%, 100,00%, 100,00%, istim redom) u odnosu na društva iz trmki gde je procenat
ovih virusa iznosio 33,33% za svaki virus. TakoĊe, sva komercijalno gajena društva bila
su inficirana makar jednim od praćenih patogena, za razliku od tradicionalno gajenih
pĉela u trmkama meĊu kojima je 66,66% bilo bez patogena.
Rezultati su otkrili znaĉajne razlike u aktivnosti CAT, GST i SOD (p<0,01) i
koncentracije MDA (p<0,002) izmeĊu komercijalnih i tradicionalnih društava što vodi
zakljuĉku da u društvima gajenim u trmkama postoji manji oksidativni stres što je
rezultiralo i manjom zastupljenošću svih praćenih patogena. U uzorcima komercijalno
gajenih pĉela zastupljenost parazita L. passim i N. ceranae bila je znaĉajno veća
(p<0,05; p<0,01, respektivno) u odnosu na uzorke iz tradicionalnih košnica, dok C.
mellificae i N. apis nisu detektovane ni u jednom uzorku. Treba istaći, da je prvi put
detektovana tripanozoma L. passim u leglu iz obe grupe ispitivanih košnica,
komercijalnih i tradicionalnih, pri ĉemu je njena zastupljenost u leglu znaĉajno manja
(p<0,01) nego u odraslim pĉelama u komercijalnim društvima, dok se kod tradicionalno
gajenih pĉela zastupljenost ove tripanozome izmeĊu odraslih pĉela i legla nije znaĉajno
razlikovala. Kod komercijalno gajenih pĉela utvrĊen je znaĉajno veći nivo iRNK za gen
GOX (p<0,01) u odnosu na tradicionalno gajena društva, što je verovatno posledica
pojaĉane potrebe prvopomenutih da ojaĉaju socijalni imunitet.
Komercijalna društva su u odnosu na tradicionalno gajena bila pod većim
oksidativnim stresom, a imala su i veću opterećenost pĉelinjim patogenima, kao i veći
nivo transkirpcije GOX gena, što je verovatno posledica ĉestih pĉelarskih manipulacija i
uznemiravanja komercijalnim društvima, kao i njihovo forsirano ekonomsko
iskorišćavanje. Sve to vodi zakljuĉku da populacija pĉela koja se uzgaja na
tradicionalan ima veći kapacitet samoodrţanja i otpornija na pĉelinje patogene,
energetski (i oksidativni) stres, a da antropogeni faktori, odnosno pĉelarski postupci,
imaju negativan uticaj na zdravlje komercijalno gajenih pĉela., Pests and pathogens are the most common causes of honey bee colony losses. American
foulbrood (AFB) is a disease of honey bee brood, caused by the bacterium
Paenibacillus larvae, White (1906), can be considered a major threat to bee health, as it
is a panzootic disease that spreads very quickly, not only from hive to hive, from apiary
to apiary, from region to region, but also from the state to the state and beyond.
European foulbrood is a bacterial disease of honey bee brood caused by several
bacterial species dominated by Melissococcus plutonius. This disease is widespread and
poses a major problem in beekeeping by reducing the production results of honey bee
colonies. Chalkbrood disease (CHB) is an infection caused by the fungus Ascosphaera
apis causing losses of commercially reared bee colonies, especially in combination with
microsporidia species of the genus Nosema sp. and a Sacbrood virus (SBV). Nosemosis
is the most common disease among adult bees caused by Nosema apis and N. ceranae,
and often leads to economic losses in beekeeping. Viral infections of bees, which have
long since reached panzootic proportions, endanger the health of bees and present
ultimate executors of bee colonies. Varroa destructor mite is the main vector of almost
all, especially viral infections, which have become a serious problem not only for
commercially reared, but also for honey bees in the wild, thanks to the mite V.
destructor as a physical and biological vector. It is clear that the health status of
colonies in commercial beekeeping is exposed to great risks, and their examination and
treatment represent a great challenge for both researchers and beekeepers.
Diseases of adult bees and bee brood in commercial beekeeping are most often a
consequence of energetic stress, caused by inadequate feeding of bee colonies, primarily
by excessive use of sugar syrup. The basis of energetic stress is oxidative stress of bee
colonies, which can be defined as an imbalance between oxidants and antioxidants in
favor of the former. Reactive oxygen species (ROS) negatively affect cellular functions
and are generated during oxido-reduction metabolic processes in the cell. They are
involved in the regulation of various mechanisms, intercellular signaling, and exert
bactericidal activity.
Honey bees, like other animals, have developed a number of enzymatic mechanisms
that resist oxidative stress, removing free radicals. An important role in these processes
is played by: superoxide dismutase (SOD), catalase (CAT) in peroxisomes, ascorbate
peroxidase, glutathione S transferase (GST) peroxidase, and the thyrodoxin /
thyroreductase system. Immunity of bees, as well as immunity in general, includes a
complex system that aims to ensure the preservation of health and survival of the
organism based on many defense mechanisms against pathogens and other harmful
noxa (physical and chemical). The honey bee achieves its social immunity with the help
of behavioral mechanisms that help the community defend itself against pathogens.
Investing in individual immunity has a high material and energy price for society, so the
association of bees (development of social structure) established joint social defense
mechanisms that enable bee society to fight more efficiently and economically against
environmental agents. The enzyme glucose oxidase (GOX) is important for the social
immunity of bees. This enzyme is a product of bee exocrine glands, and its role is in the
"conservation" of bee products, honey and bee bread, thus preventing their spoilage.
GOX participates in the process of catalysis of -D glucose to gluconic acid and
hydrogen peroxide. Hydrogen peroxide has antiseptic properties, thus contributing to
additional protection and social immunity of bees.
The aim of the research was to determine: whether there are differences between
traditional and modern bee colonies in terms of the presence of pathogens and the
occurrence of bee brood diseases, with the analysis of biological (expression level of the
GOX gene and oxidative stress factors) and anthropogenic factors (beekeeping
management) on monitored bee brood and adult bee diseases. The field part of the
research was performed in the area of the Peshterska plateau (Sjenica municipality,
Serbia) on 144 asymptomatic bee communities. Samples of adult bees and bee brood
were taken from modern (DB) hives and traditional "trmka" hives to determine the
presence of brood pathogens (Paenibacilus larvae, Melissococus plutonius, Ascosphera
apis, Sacbrood virus, trypanosomatids Lotmaria passim and Crithidia mellificae) and
adult bees (acute bee paralysis virus, chronic bee paralysis virus, deformed bee wings
virus, trypanosomatids Lotmaria passim and Crithidia mellificae, microsporidia
Nosema apis and Nosema ceranae) as well as for determination of oxidative stress
levels by analysis of activity of enzymes: SOD, CAT, GST and concentration of
malondialdehyde (MDA). From the same hives, samples were taken to estimate the
expression the GOX gene as a parameter of social immunity. Pathogen detection was
performed by bacterial isolation as well as using PCR and real-time PCR, in accordance
with the OIE standards. The results showed that among commercially reared colonies,
P. larvae, was represented in 16.67% of samples, A. apis in 15.83%, while SBV was
detected in 96.67% of samples. However, in the brood from colonies raised in
traditional hives, only SBV in the percentage of 33.33% of samples was found. Further
research found that in adult bees from commercially reared colonies, viruses ABPV,
CBPV and DWV (83.33%, 100.00%, 100.00%, respectively), were significantly (p
<0.001) more represented compared to adult bees from „trmka― hives, where the
percentage of these viruses was 33.33% for each. Also, all commercially reared colonies
were infected with at least one of the monitored pathogens, in contrast to traditionally
reared bees, of which 66.66% were pathogen-free.
The results revealed significant differences of activities of CAT, GST and SOD
(p<0.01) and MDA concentration (p<0.002) between commercial and traditional
colonies, which leads to the conclusion that colonies grown in „trmka― hives had less
oxidative stress level, which resulted in lower presence of all monitored pathogens.
In samples of bees from commercially bred colonies, the prevalence of parasites L.
passim and N. ceranae was significantly higher (p<0.05; p<0.01, respectively)
compared to samples from traditional hives, while C. mellificae and N. apis were not
detected in any sample. It should be noted that L. passim was detected for the first time
in a brood from both groups of examined hives, commercial and traditional, with its
presence in the brood significantly lower (p<0.01) than in adult bees in commercial
colonies, while in traditionally reared bees, the presence of this trypanosome did not
differ significantly between adult bees and brood. In commercially bred bees, a
significantly higher level of mRNA for the GOX gene was found (p<0.01) compared to
traditionally bred colonies, which is probably a consequence of the increased need of
the former to strengthen social immunity. Compared to traditional beekeeping,
commercial colonies were under greater oxidative stress, and had a higher load of bee
pathogens as well as a higher level of transcription for the GOX gene, which is probably
due to frequent beekeeping and harassment of commercial colonies, as well as their
forced economic exploitation. All this leads to the conclusion that the population of
bees raised in the traditional way is more self-sustaining and resistant to bee pathogens,
energetic (and oxidative) stress, and that the anthropogenic factors (beekeeping
practices) have a negative impact on the health of commercially reared bees.",
publisher = "Beograd : Fakultet veterinarske medicine",
journal = "2022",
title = "Komparativno ispitivanje prisustva uzročnika i bolesti pčelinjeg legla tradicionalno i savremeno gajenih pčelinjih društava uz analizu uticaja nekih bioloških i antropogenih faktora",
url = "https://hdl.handle.net/21.15107/rcub_veterinar_3360"
}
