UNIVERSITY OF MALTA
LIFE SCIENCE RESEARCH SEMINARS
Web: http://www.um.edu.mt/events/scisem/
Email: scisem@um.edu.mt
Abstract form
Title: Determination of the genetic status of the local honey bee Apis
mellifera ruttneri
Presenter: Sheryl Sammut
Contact address: Dawl Zghir, Triq il- Ballut, Attard
Tel: 21434541
Fax: /
Email: sheryl.sammut@gmail.com
Presentation date: 26/05/2014
Abstract
Bees are essential to the ecosystem, pollinating a great variety of both insect pollinated, wild plants
and crops. Rampant importation of other honey bee subspecies in the Maltese Islands, is endangering
the unique genetic identity of the Maltese subspecies, Apis mellifera ruttneri. To date only one study
(Sheppard et al., 1997) has been published locally concerning the genetic identity of this important
species. Therefore, the main goal of this project is to investigate and determine the status of the native
honey bee in the present time.
Between October and December 2013, specimens of Apis mellifera were collected from a number of
apiaries found in the Maltese Islands. These were subjected to morphometric analysis, where worker
bees were dissected and examined for the 35 morphometric parameters proposed by Ruttner., et al
(1978), as well as mitochondrial DNA analysis. In the latter, the total DNA was extracted using a
modified CTAB extraction method (Doyle & Doyle, 1987), after which DNA analysis was performed on
the intergenic region between the Cytochrome oxidase C sub-unit I (COI) and Cytochrome oxidase C
sub-unit II (COII) genes. The latter intergenic region spans a tRNAleu gene as well as repeats of noncoding sequences referred to as P and Q sequences. The mentioned intergenic region was amplified
using the primer pair E2-H2. Amplified fragments were monitored by gel electrophoresis using
agarose and PAGE, followed by RFLP analysis using the restriction endonuclease Dra1. A sample of
each undigested product, for every specimen, was sequenced by Sanger sequencing.
The samples were found to originate from three main evolutionary lineages; M lineage (northern and
western Europe), A lineage (Africa) and C lineage (southeastern Europe). 80% of the 106 samples
analysed for the COI-COII region pertained to different A lineages (74% A4 and 6% A1), 10%
belonged to the C1 lineage, 8% were part of the C2 lineage, whilst 2% were found to the part of the
M4 lineage. The intergenic region between the tRNAlue gene and the COII of the A and M lineages
varied in the composition of the P and Q sequences and further details will be presented in the
presentation.
In contrast to other studies abroad, where the Dra1 restriction site in A, C and M lineages is always
present, only 43% of the analysed samples locally showed restriction sites for Dra1 at the COI-COII
intergenic region. While all samples of C1, C2 and M4 origin contained the restriction site, only 20% of
the A lineage samples showed restriction sites for Dra1. Evidence is presented that this smaller pool
of Apis mellifera may be remnants of the native A.m. ruttneri as originally described by Sheppard et al
in 1997.
Multiple alignments and phylogenetic analysis using neighbor-joining methods were performed, where
the C, A and M4 lineages were clearly distinct clusters. With respect to morphometric analysis,
multivariate analysis, including Principal Components Analysis (PCA), is currently being carried out.