PCRmax Naegleria (all species) Instruction Manual

18S ribosomal RNA (18S) gene

Naegleria species

PCRmax Ltd qPCR test

TM

150 tests

For general laboratory and research use only

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Naegleria is a genus of the Vahlkampfiidae family containing many species, of which only
one is known to infect humans; Naegleria fowleri. Species of the Nagleria genus have a life
cycle with three stages: cysts; trophozoites which replicate by promitosis where the nuclear
membrane remains intact and flagellated bodies. Infection with N. fowleri causes the rare
but fatal disease primary amoebic encephalitis.

Cysts can be found in warm fresh-water sources and transform into trophozoites which
after inhalation, cross the olfactory neuroepithelium in the nasal cavity and move to the
brain where they cause encephalitis. Initial trophozoite infection causes necrosis of the
olfactory bulbs due to promitotic replication. After replication the infection spreads to the
rest of the cerebellum eventually resulting in fatality.

Within 3-7 days of infection, symptoms including olfactory dysfunction and loss of smell are
present as the olfactory bulb is destroyed. As the infection spreads, symptoms become
more generalized and include nausea, headache and seizures leading to irreversible
coma. Infected individuals most commonly die from respiratory failure as the infection
spreads to the brain stem after around two weeks.

Antimicrobial agents are used to treat suspected Naegleria infections but even with this
treatment, the fatality rate is around 97%. Treatment success is low partly due to the late
use of these microbial agents as the initial symptoms are difficult to diagnose but also due
the increased rapidity of the infection during later stages.

Introduction to Naegleria species

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MIN

MAX

Our kit for Naegleria_spp has been designed for the specific and exclusive in vitro
quantification of this genus . The 18S ribosomal gene, is the ideal target to achieve a
broad based detection profile for all species within this genus. The primers and probe
sequences in this kit have 100% homology with a broad range of clinically relevant
reference sequences based on a comprehensive bioinformatics analysis. Other closely
related species are not detected.

The PCR Max qPCR Kit for Naegleria species (Naegleria_spp) genomes is designed for
the in vitro quantification of Naegleria_spp genomes. The kit is designed to have the
broadest detection profile possible whilst remaining specific to the Naegleria_spp genome.

The primers and probe sequences in this kit have 100% homology with a broad range of
Naegleria_spp sequences based on a comprehensive bioinformatics analysis.

If you require further information, or have a specific question about the detection profile of
this kit then please send an e.mail to [e-mail] and our bioinformatics team will
answer your question.

Specificity

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Kit Contents

• Naegleria_spp specific primer/probe mix (150 reactions BROWN)

FAM labelled

• Naegleria_spp positive control template (for Standard curve RED)

• Internal extraction control primer/probe mix (150 reactions BROWN)

VIC labelled as standard

• Internal extraction control DNA (150 reactions BLUE)

• Endogenous control primer/probe mix (150 reactions BROWN)

FAM labelled

• RNAse/DNAse free water (WHITE)

for resuspension of primer/probe mixes and internal extraction control DNA

• Template preparation buffer (YELLOW)

for resuspension of positive control template and standard curve preparation

Reagents and equipment to be supplied by the user

Real-Time PCR Instrument

DNA extraction kit

This kit designed to work well with all processes that yield high quality DNA with minimal
PCR inhibitors.

Lyophilised 2x qPCR Mastermix

This kit is designed to be compatible with all commercially available Mastermixes that run
with standard cycling conditions.

Pipettors and Tips

Vortex and centrifuge

Thin walled 1.5 ml PCR reaction tubes

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