PMID- 30470185
OWN - NLM
STAT- MEDLINE
DCOM- 20190705
LR  - 20240214
IS  - 1471-2180 (Electronic)
IS  - 1471-2180 (Linking)
VI  - 18
IP  - Suppl 1
DP  - 2018 Nov 23
TI  - High-sensitivity detection of cryptic Wolbachia in the African tsetse fly 
      (Glossina spp.).
PG  - 140
LID - 10.1186/s12866-018-1291-8 [doi]
LID - 140
AB  - BACKGROUND: In African tsetse flies Glossina, spp. detection of bacterial 
      symbionts such as Wolbachia is challenging since their prevalence and 
      distribution are patchy, and natural symbiont titers can range at levels far 
      below detection limit of standard molecular techniques. Reliable estimation of 
      symbiont infection frequency, especially with regard to interrelations between 
      symbionts and their potential impact on host biology, is of pivotal interest in 
      the context of future applications for the control and eradication of 
      Glossina-vectored African trypanosomosis. The presence or absence of symbionts is 
      routinely screened with endpoint polymerase chain reaction (PCR), which has 
      numerous advantages, but reaches its limits, when detecting infections at natural 
      low titer. To not only determine presence of native tsetse symbionts but also to 
      localize them to specific host tissues, fluorescence in situ hybridization (FISH) 
      can be applied. However, classic FISH assays may not detect low-titer infections 
      due to limitations in sensitivity. RESULTS: We have compared classic endpoint PCR 
      with high-sensitivity blot-PCR. We demonstrate that the latter technique allows 
      for clear detection of low-titer Wolbachia in the morsitans and palpalis groups 
      while classic endpoint PCR does not. In order to localize Wolbachia in situ in 
      high and low-titer Glossina species, we applied high-end Stellaris(R) rRNA-FISH. We 
      show that with this high sensitivity method, even low amounts of Wolbachia can be 
      traced in specific tissues. Furthermore, we highlight that more tissues and 
      organs than previously recorded are infested with Wolbachia in subspecies of the 
      morsitans and palpalis groups. CONCLUSIONS: Our results demonstrate that overall 
      symbiont infection frequencies as well as the presence in specific host tissues 
      may be underestimated when using low-sensitivity methods. To better understand 
      the complex interrelation of tsetse flies and their native symbionts plus the 
      pathogenic trypanosomes, it is important to consider application of a broader 
      range of high-sensitivity detection tools.
FAU - Schneider, Daniela I
AU  - Schneider DI
AD  - Department Cell and Developmental Biology, Center for Anatomy and Cell Biology, 
      Medical University of Vienna, Vienna, Austria.
AD  - Present Address: Department of Epidemiology of Microbial Diseases, Yale School of 
      Public Health, New Haven, CT, USA.
FAU - Parker, Andrew G
AU  - Parker AG
AD  - Insect Pest Control Laboratory, Joint FAO/IAEA Division of Nuclear Techniques in 
      Food and Agriculture, Vienna, Austria.
FAU - Abd-Alla, Adly M
AU  - Abd-Alla AM
AD  - Insect Pest Control Laboratory, Joint FAO/IAEA Division of Nuclear Techniques in 
      Food and Agriculture, Vienna, Austria.
FAU - Miller, Wolfgang J
AU  - Miller WJ
AD  - Department Cell and Developmental Biology, Center for Anatomy and Cell Biology, 
      Medical University of Vienna, Vienna, Austria. wolfgang.miller@meduniwien.ac.at.
LA  - eng
GR  - P 22634/FWF_/Austrian Science Fund FWF/Austria
GR  - P 28255/FWF_/Austrian Science Fund FWF/Austria
PT  - Comparative Study
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20181123
PL  - England
TA  - BMC Microbiol
JT  - BMC microbiology
JID - 100966981
RN  - 0 (Bacterial Outer Membrane Proteins)
SB  - IM
MH  - Animals
MH  - Bacterial Outer Membrane Proteins/genetics
MH  - Female
MH  - In Situ Hybridization, Fluorescence/*methods
MH  - Insect Vectors/microbiology
MH  - Limit of Detection
MH  - Male
MH  - Polymerase Chain Reaction/*methods
MH  - Sensitivity and Specificity
MH  - Symbiosis
MH  - Tsetse Flies/*microbiology
MH  - Wolbachia/genetics/*isolation & purification
PMC - PMC6251158
OTO - NOTNLM
OT  - Low-titer symbiont detection limit
OT  - Stellaris(R) fluorescence in situ hybridization
OT  - Tissue tropism
OT  - Wolbachia
COIS- ETHICS APPROVAL AND CONSENT TO PARTICIPATE: Not applicable. CONSENT FOR 
      PUBLICATION: Not applicable. COMPETING INTERESTS: The authors declare that they 
      have no competing interests. PUBLISHER'S NOTE: Springer Nature remains neutral 
      with regard to jurisdictional claims in published maps and institutional 
      affiliations.
EDAT- 2018/11/25 06:00
MHDA- 2019/07/06 06:00
PMCR- 2018/11/23
CRDT- 2018/11/25 06:00
PHST- 2018/11/25 06:00 [entrez]
PHST- 2018/11/25 06:00 [pubmed]
PHST- 2019/07/06 06:00 [medline]
PHST- 2018/11/23 00:00 [pmc-release]
AID - 10.1186/s12866-018-1291-8 [pii]
AID - 1291 [pii]
AID - 10.1186/s12866-018-1291-8 [doi]
PST - epublish
SO  - BMC Microbiol. 2018 Nov 23;18(Suppl 1):140. doi: 10.1186/s12866-018-1291-8.