Eugenia Soboleva presents
CHARACTERISTICS OF THE INVERSION POLYMORPHISM OF THE NATURAL POPULATIONS OF SIBERIA OF MALARIA MOSQUITOES ANOPHELES BEKLEMISHEVI USING FLUORESCENT IN SITU HYBRIDIZATION
Laboratory of the ecology, genetic and environmental protection
National Research Tomsk state university; Tomsk (Russia)
INTRODUCTION
Anopheles beklemishevi Stegnii et Kabanova is a malaria mosquito and member of the Maculipennis group inhabiting in Palearctic. Its distribution area is the northernmost among malaria mosquitoes of Eurasia and covers the harshest climatic zones of Western and Eastern Siberia as well as northern Europe. The genetic mechanisms of cold resistance of Anopheles mosquitoes may be related to adaptive inversion polymorphism. An. beklemishevi is a one of the rare species of malaria mosquitoes, in which inversions in X chromosome are widespread in natural populations. Until now polymorphic inversions X1 and X2 have been found, according to the literature and our own observations, only in heterozygous state. This could be seen as a Hardy-Weinberg disequilibrium. Because of peculiarities of localization of inversions, the analysis of the order of chromosomal banding of polytene chromosomes does not allow reliably detect inverted homozygous.

The aim of our study
*MD - microdissection FISH - fluorescence in situ hybridization
The aim of this study was to determine the frequencies of homozygotes for inversions in the X chromosome in natural populations of An. beklemishevi using the MD-FISH* method
read more - A standard photomap of ovarian nurse cell chromosomes and inversion polymorphism in Anopheles beklemishevi
What have we done?
Using the microdissection and PCR we developed a fluorescence DNA-probe marking 1a-3a region of standard X chromosome (X0) of An. beklemishevi. This region includes only one, distal, breakpoint of polymorphic inversions X1 and X2. After FISH with a standard X chromosome the signal of DNA-probe appears to be solid, however in inverted variants of chromosome X1 and X2 the labeled region divided into two parts, in a specific way for each inversion which makes it easy to detect the genotype
FISH result
Fluorescence in situ hybridization of 1A-3B DNA-zond with X0 (a), X1 (b) and X2 (c) chromosomes of An. beklemishevi. CC - chromocenter.
Scheme of microdissection of a part of the sex chromosome of Anopheles beklemishevi
Microdissection of the 1A-3B region of the X chromosome of Anopheles beklemishevi. Region 1A-3B is arrowed before microdissection on A and the place where it was on B, when microdissection has performed.
Results
we found something that no one else has found
The material for the study were samples of Anopheles mosquitoes larvae had been caught in natural reservoirs in Siberia during the period from 2019 to 2021. The geography of samples includes specimens from Yamalo-Nenets Autonomous Okrug (YaNAO), Khanty-Mansi Autonomous Okrug (KhMAO), Krasnoyarsk Krai (KK), Tomsk Oblast (TO) and Altay Republic.

As the result, it has been identified 480 larvae of An. beklemishevi from 8 natural populations. The distance between the northernmost (village Shuryshkary) and southernmost one (village Ozernoye) was almost 2000 kilometers in a straight line. In a car this way will be approximately 3100 kilometers. Usually, the larvae of An. beklemishevi inhabits with An. messeae and An. daciae together, but in the north and in the highlands as in the early-summer period, mainly individuals only of this species can be found.


The analysis of inversion polymorphism in Western Siberia
The standard homozygous cytotype was found in all populations with clear predominance. Heterozygous X02 variant was found in most populations with the frequencies from 3.6 % to 28.6 %, but variant X01 was not found at all. Also, we found for the first time X2 cytotype in homozygous in six populations with the frequencies from 3.2% to 17.4%, while homozygous individuals with X1 inversion in homozygous state with the frequencies from 3.6 % to 3.9 % in only two populations.
This research showed the effectiveness of using MD-FISH method to detect homozygotes for inversions in the X chromosome An. beklemishevi which can occur with high frequency.

The study was supported by RSF grant №21-14-00182

Laboratory of the ecology, genetic
and environmental protection
Institute of biology of Tomsk state university
Meet Our Team
The smartest people work every day to make science great again!
  • Gleb Artemov
    PhD of biology
    Acting laboratory manager; search and solution of scientific problems
  • Alina Kohkanenko
    PhD of biology
    Manager of syntetic biology division, specialist of Oxford Nanopore sequencing
  • Vladimir Burlak
    PhD of biology
    The main parasitologist in team, studies the spread of dirofilariasis in Siberia
  • Vladimir Stegnii
    Doctor of Biological Science
    The founder of laboratory and mosquito-school in Siberia
  • Eugenia Soboleva
    PhD student
    Studies inversion polymorphism of Anopheles mosquito in Siberia
  • Mohamed Kader
    Scientist
    Studies dirofilariasis of Anopheles mosquito in Siberia
  • Anna Kirilenko
    PhD student
    Synthetic biology division developer
  • Tatyana Babai
    Scientist
    Synthetic biology division developer
  • Kirill Kirilenko
    PhD student
    Bioinformatic division, mobile elements catcher, genome assembly
  • Svetlana Alexeeva
    Scientist
    Non-malaria mosquito specialist, Sanger sequencing specialist
  • Tatyana Ananina
    PhD of biology
    Manager of Cell biology division, Diptera oogenesis specialist
  • Ximena Calderon
    PhD student
    Rearing Anopheles mosquitoes, geomorphometric analysis
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