<div dir="ltr">Dear J-C,<div><br></div><div>Thank you so much for the pdf reprint of hsp70.</div><div><br></div><div> It is great to see that the use of "clinical samples" was mentioned in the Materials and Methods, although I didn't find further reference to them in the Results and Discussion in that article. I may have missed ? As you know, I feel very strongly that we ought to examine amastigotes directly in the clinical samples instead of or in addition to cultured promastigotes. I gather from your publications that the latter may or may not be the actual causative agent of the disease seen in the patient of concern. We have preliminary data, which strongly support your finding. </div>
<div><br></div><div>As I have also mentioned that ultimately we will have to do genomic sequencing of individual amastigotes from the clinical samples. Your group has been in active pursue of genomics, transcriptomics and proteomics in a number of different context. You are undoubtedly knowledgeable and fully aware of the importance of such approaches for meta-analysis. </div>
<div><br></div><div>PCR amplification of individual genes for RFLP and sequencing analysis certainly has its merit, but this is an interim approach, facing the difficulty of specificity versus sensitvity in selecting multicopy vs single-copy marker genes and host-parasite sequence overlaps (when using clinical samples). We have discussed this and other issues in our article published in 2007 on the 1.4 kb<i> nagt</i> that was PCR-amplified for sequnece and RFLP analyses from hundreds of promastigote cultures obtained by ourselves or generously providfed by colleagues mainly in the Eurasian continents. The outcome was verified by similar analysis of 4 addtional single-copy genes. Still, the sample is too small in size and biased by using cultured promastigotes, as stated in the 1st paragraph of the Discussion section. Toward the end, we concluded that genomic sequencing will be necessary.</div>
<div><br></div><div>During the interim period, the best approach is to PCR-amplify multiple DNAs in different genetic sites of amastigotes direclty from clinical samples for sequence and RFLP analysis, including the hsp70 presented in your article. </div>
<div><br></div><div>KP </div></div><div class="gmail_extra"><br><br><div class="gmail_quote">On Sat, Nov 2, 2013 at 2:28 AM, Jean-Claude Dujardin <span dir="ltr"><<a href="mailto:JCDujardin@itg.be" target="_blank">JCDujardin@itg.be</a>></span> wrote:<br>
<blockquote class="gmail_quote" style="margin:0 0 0 .8ex;border-left:1px #ccc solid;padding-left:1ex">
<div lang="NL-BE" link="blue" vlink="purple">
<div>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">Dear KP, dear all, can I also recommend hsp70 PCR (followed by RFLP or sequencing)? The same primers function worldwide for all Leishmania species
and allow discrimination of Ldonovani from Linfantum. Sensitivity in clinical samples is very good. See attached one of the paper of our team; if you want more details ask Gert Van der Auwera (in cc)</span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">Best regards</span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">JC</span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d"> </span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">Follow me at
</span><span style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d"><a href="https://twitter.com/DujardinBiomed" target="_blank"><span lang="EN-US" style="color:blue">https://twitter.com/DujardinBiomed</span></a></span><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d"></span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d"> </span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">Prof Dr Jean-Claude Dujardin</span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">Institute of Tropical Medicine</span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">Dept of Biomedical Sciences, Head</span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">Molecular Parasitology Unit, Head</span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">Nationalestraat, 155</span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">B-2000 Antwerpen</span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">Belgium</span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d"> </span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d"><a href="tel:32.3.2476355" value="+13232476355" target="_blank">32.3.2476355</a></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d"> </span></p>
<p class="MsoNormal"><b><span lang="EN-US" style="font-size:10.0pt;font-family:"Tahoma","sans-serif"">From:</span></b><span lang="EN-US" style="font-size:10.0pt;font-family:"Tahoma","sans-serif""> <a href="mailto:leish-l-bounces@lineu.icb.usp.br" target="_blank">leish-l-bounces@lineu.icb.usp.br</a> [mailto:<a href="mailto:leish-l-bounces@lineu.icb.usp.br" target="_blank">leish-l-bounces@lineu.icb.usp.br</a>]
<b>On Behalf Of </b>Kwang-Poo Chang<br>
<b>Sent:</b> vrijdag 1 november 2013 17:59<br>
<b>To:</b> Σωτηριάδου Καίτη<br>
<b>Cc:</b> Lawyer, Phillip (NIH/NIAID) [E]; <a href="mailto:leish-l-bounces@lineu.icb.usp.br" target="_blank">leish-l-bounces@lineu.icb.usp.br</a>; Rossella Colomba Lelli; Leish-L; <a href="mailto:caporalevincenzo@gmail.com" target="_blank">caporalevincenzo@gmail.com</a></span></p>
<div><div class="h5"><br>
<b>Subject:</b> Re: [Leish-l] why is Leishmania donovani restricted to humans inIndian subcontinent?</div></div><p></p><div><div class="h5">
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<p class="MsoNormal">Dear Ketty,</p>
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<p class="MsoNormal">Thanks a lot for the information ! </p>
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<p class="MsoNormal">Could you kindly send the pdf reprints for k26, CBP and hsp90 PCR for diagnostic discrimination of L donovani complex for clinical samples ? I assume that they work for dog, sanfly and human samples ?</p>
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<p class="MsoNormal">KP</p>
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<p class="MsoNormal">On Fri, Nov 1, 2013 at 5:34 AM, Σωτηριάδου Καίτη <<a href="mailto:ksoteriadou@pasteur.gr" target="_blank">ksoteriadou@pasteur.gr</a>> wrote:</p>
<div>
<div>
<div>
<p class="MsoNormal"> </p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">Hi! In our hands also 13A/13B kDNA had the same problems. We came to the conclusion that the balance sensitivity/specificity is not good. Specifity/contamination
is an issue in that sense.</span><span lang="EN-US" style="color:#1f497d"></span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">If you want to discriminate species belonging to the L. donovani complex k26-PCR, CPB and hsp90 give the same results that are not always in
agreement with MLEE. ITS (among others) is an excellent tool for discriminating species causing VL or CL in Middle East. Please note that the latter tools can be applied on clinical samples which is an important issue. I could sent you the relevant papers
in a pdf format or the citation, there are several. Please let me know.</span><span lang="EN-US" style="color:#1f497d"></span></p>
<p class="MsoNormal"><span lang="EN-US" style="font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1f497d">Ketty Soteriadou (Greece)</span><span lang="EN-US" style="color:#1f497d"></span></p>
<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif""> </span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">The one by Reale is the 13A/13B kDNA protocol. Its extremely sensitive. We've tried it. The problem is that after sometime, one start to see that its becoming positive for all samples.
Then we gave it up for the benefit of genomic-based methods. Perhaps we should have used UNG-uracil in the PCR setup as an anti-contamination agent.</span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif""> </span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">Amer</span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">Palestine</span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif""> </span></p>
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<hr size="1" width="100%" align="center">
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<p class="MsoNormal"><b><span style="font-family:"Arial","sans-serif"">From:</span></b><span style="font-family:"Arial","sans-serif""> Stefano Reale <<a href="mailto:stefano.reale@izssicilia.it" target="_blank">stefano.reale@izssicilia.it</a>><br>
<b>To:</b> Kwang-Poo Chang <<a href="mailto:kwangpoo.chang@rosalindfranklin.edu" target="_blank">kwangpoo.chang@rosalindfranklin.edu</a>>
<br>
<b>Cc:</b> "Lawyer, Phillip (NIH/NIAID) [E]" <<a href="mailto:PhillipL@niaid.nih.gov" target="_blank">PhillipL@niaid.nih.gov</a>>; "<a href="mailto:leish-l-bounces@lineu.icb.usp.br" target="_blank">leish-l-bounces@lineu.icb.usp.br</a>" <<a href="mailto:leish-l-bounces@lineu.icb.usp.br" target="_blank">leish-l-bounces@lineu.icb.usp.br</a>>;
Rossella Colomba Lelli <<a href="mailto:rossella.lelli@izssicilia.it" target="_blank">rossella.lelli@izssicilia.it</a>>; Leish-L <<a href="mailto:Leish-L@lineu.icb.usp.br" target="_blank">Leish-L@lineu.icb.usp.br</a>>; "<a href="mailto:caporalevincenzo@gmail.com" target="_blank">caporalevincenzo@gmail.com</a>"
<<a href="mailto:caporalevincenzo@gmail.com" target="_blank">caporalevincenzo@gmail.com</a>>
<br>
<b>Sent:</b> Thursday, October 31, 2013 8:19 AM<br>
<b>Subject:</b> Re: [Leish-l] why is Leishmania donovani restricted to humans in Indian subcontinent?</span><span style="font-family:"Helvetica","sans-serif""></span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">Hallo Im one of the author of this PCR protocol. I optimised it by specific primers and probe for infantum. It work very well and I showed the Lod and Loq in variuos biological material
taken from leishmaniotic dogs. In my lab in Palermo I have a number if collected strains stored in liquid N2 ready to create the DNA standard referement serial dilutions fir Real time test. I could help who should have these DNA.<br>
Best regards</span></p>
</div>
<div>
<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">Il giorno 31/ott/2013 00:07, "Kwang-Poo Chang" <<a href="mailto:kwangpoo.chang@rosalindfranklin.edu" target="_blank">kwangpoo.chang@rosalindfranklin.edu</a>> ha scritto:</span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">Dear Carlos,
</span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif""> </span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">One of my colleagues in Naples, Italy has used primer set for kinetoplast minicircle conserved region, Considering ~10,000 copies of minicircvles per Leishmania, this is probably the most
sensitive PCR, although the primer set can also amply all trypanosomes or trypanosomatid protozoa (including Leptomonas reported to exist in the Indian kala-azar splenic aspirates). Specificity could be an issue in that sense.</span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">KP </span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">On Thu, Oct 10, 2013 at 7:31 PM, Carlos Lobo <<a href="mailto:carloshlobo@gmail.com" target="_blank">carloshlobo@gmail.com</a>> wrote:</span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">Hello guys, good night.</span></p>
</div>
<div>
<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">I'm taking advantage of this opportunity to exchange scientific information, much of which I have learned to ask for help.<br>
I'll start a project on canine leishmaniasis in northeastern Brazil and I need to diagnose, by PCR, dogs infected and not infected with Leish.<br>
I've been doing some analysis of PCR for other experiments, but never did for Leish, could someone give me some tips? Like which primer to use? Should I collect blood or tissue samples to have more reliability?<br>
Thank you.<br>
Carlos Henrique</span></p>
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<p class="MsoNormal" style="margin-bottom:12.0pt"><span style="font-family:"Helvetica","sans-serif""> </span></p>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">2013/9/23 Tamrat Abebe Zeleke <<a href="mailto:tabebezeleke@gmail.com" target="_blank">tabebezeleke@gmail.com</a>></span></p>
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<p class="MsoNormal" style="margin-bottom:12.0pt"><span style="font-family:"Georgia","serif"">Dear Carlos,</span></p>
</div>
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<div>
<p class="MsoNormal"><span style="font-family:"Georgia","serif"">I agree with the comment given by Phillip Lawyer. The molecular analysis of strains or isolates from India, Kenya, and South Western Ethiopia also supports this notion. However, the issue of distinct
strains in Sudan and North Ethiopia opts for the fact that East Africa may be the origin of at least the naughty L. donovani strains circulating in the region.
</span></p>
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</span></p>
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<p class="MsoNormal"><span style="font-family:"Tahoma","sans-serif";color:#6fa8dc">Tamrat Abebe</span><span style="font-family:"Helvetica","sans-serif""></span></p>
</div>
<p class="MsoNormal"><span style="font-family:"Tahoma","sans-serif";color:#6fa8dc">Addis Ababa University College of Health Sciences, School of Medicine ,<br>
Department of Microbiology, Immunology & Parasitology </span><span style="font-family:"Helvetica","sans-serif"">
</span></p>
<div>
<p class="MsoNormal"><span style="font-family:"Tahoma","sans-serif";color:#6fa8dc">Tikur Anbessa Hospital</span><span style="font-family:"Helvetica","sans-serif""></span></p>
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<p class="MsoNormal"><span style="font-family:"Tahoma","sans-serif";color:#6fa8dc">Second floor room number 76<br>
Tel: <a href="tel:%2B251%20911%20447227" value="+251911447227" target="_blank">+251 911 447227</a>(mobile)<br>
Email:</span><span style="font-family:"Comic Sans MS";color:#6fa8dc"> </span><span style="font-family:"Helvetica","sans-serif";color:#6fa8dc"><a href="mailto:tamrat.abebe@aau.edu.et" target="_blank"><span style="font-family:"Tahoma","sans-serif"">tamrat.abebe@aau.edu.et</span></a></span><span style="font-family:"Helvetica","sans-serif"">
</span></p>
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</span></b><span style="font-family:"Helvetica","sans-serif""> <br>
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<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">On Mon, Sep 16, 2013 at 4:37 AM, Lawyer, Phillip (NIH/NIAID) [E] <<a href="mailto:PhillipL@niaid.nih.gov" target="_blank">PhillipL@niaid.nih.gov</a>> wrote:</span></p>
<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif"">Dear Carlos,<br>
<br>
<br>
<br>
For what it's worth, I believe it most likely happened the other way around: Leishmania donovani was probably introduced to East Africa from India during the late 1800s when laborers were brought from India to work in the Kenya building the railroad from Mombasa
to Uganda and on other infrastructure projects. Leishmania donovani in East Africa is manifest as kala azar and is anthroponotic, the same as in India. The main vector in Kenya is Phlebotmus martini, which tends to breed in termite mounds, often associated
with human dwellings. Other Symphlebotomus species, Ph. vansomerenae and Ph. celiae have also been implicated in L. donovani transmission.<br>
<br>
<br>
<br>
Regards,<br>
<br>
<br>
<br>
Phil Lawyer<br>
<br>
________________________________<br>
From: Carlos Brisola Marcondes [<a href="mailto:cbrisolamarcondes@gmail.com" target="_blank">cbrisolamarcondes@gmail.com</a>]<br>
Sent: Friday, August 30, 2013 8:41 AM<br>
To: Leish-L; <a href="mailto:leish-l-bounces@lineu.icb.usp.br" target="_blank">leish-l-bounces@lineu.icb.usp.br</a><br>
Subject: [Leish-l] why is Leishmania donovani restricted to humans in Indian subcontinent?</span></p>
<div>
<div>
<p class="MsoNormal"><span style="font-family:"Helvetica","sans-serif""><br>
Dear all,<br>
Leishmania donovani seems to have been introduced from East Africa to Indian subcontinent, where it has infected mostly humans, differently from East African foci.<br>
Why does this occur? Is this caused by feeding preferences of Phlebotomus argentipes, which bites mostly ruminants and humans and is associated to houses? Or are dogs rarer in that region than in Brazil, where these animals are important reservoirs of Leishmania
infantum and frequently bitten by Lutzomyia longipalpis?<br>
<br>
Sincerely yours<br>
prof. dr. Carlos Brisola Marcondes<br>
Dept. Microbiol. Imunol. Parasitol./CCB<br>
Federal University of Santa Catarina<br>
Florianópolis (SC)<br>
CV: <a href="http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4783901J2" target="_blank">
http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4783901J2</a><br>
blog: <a href="http://entomomedica.blogspot.com.br/" target="_blank">http://entomomedica.blogspot.com.br/</a><br>
<br>
<br>
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