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Increased detection of invasive enteropathogenic bacteria in pre-incubated blood culture materials by real-time PCR in comparison with automated incubation in Sub-Saharan Africa

Frickmann, Hagen,Dekker, Denise,Boahen, Kennedy,Acquah, Samuel,Sarpong, Nimako,Adu-Sarkodie, Yaw,Schwarz, Norbert G.,May, Jü,rgen,Marks, Florian,Poppert, Sven,Wiemer, Dorothea F.,Hagen, Ralf M. Informa Healthcare 2013 Scandinavian journal of infectious diseases Vol.45 No.8
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Background: Invasive enteropathogenic bacteria can cause systemic infections. Data from studies with PCR detection suggest, at least for Salmonella enterica, that blood culture may lead to underestimation in the tropics. Corresponding data are lacking for other invasive enteropathogenic bacteria. We compared classical blood culture and molecular methods for the diagnosis of blood infections. Methods: A real-time multiplex PCR for Salmonella spp., Shigella spp./entero- invasive Escherichia coli (EIEC), Yersinia spp., and Campylobacter jejuni was applied to 2321 retained blood culture samples from Ghanaian patients, after enrichment by automated culture. Results: PCR detected Salmonella DNA in 56 out of 58 pre-incubated Ghanaian blood cultures with growth of S. enterica. In 2 samples molecular diagnosis was only possible after 1:10 dilution. Twenty-two samples negative by blood culture and 1 positive with Micrococcus spp. were PCR-positive for Salmonella spp. In addition, 3 Shigella spp./EIEC, 2 Yersinia spp., and 1 C. jejuni were detected by PCR but not by culture growth. Conclusions: Real-time PCR was more sensitive in identifying invasive enteropathogenic bacteria than automated blood culture, which is hampered by a lack of evidence-based standardization of pre-analytic conditions in the tropics. Primary agar culture and Gram-staining prior to automated blood culture is advisable in cases where transportation times are long.
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Presence of Borrelia spp. DNA in ticks, but absence of Borrelia spp. and of Leptospira spp. DNA in blood of fever patients in Madagascar

Hagen, Ralf Matthias,Frickmann, Hagen,Ehlers, Julian,Krü,ger, Andreas,Margos, Gabriele,Hizo-Teufel, Cecilia,Fingerle, Volker,Rakotozandrindrainy, Raphael,Kalckreuth, Vera von,Im, Justin,Pak, Gi De Elsevier 2018 Acta tropica Vol.177 No.-
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Abstract The occurrence of tick-borne relapsing fever and leptospirosis in humans in Madagascar remains unclear despite the presence of their potential vectors and reservoir hosts. We screened 255 Amblyomma variegatum ticks and 148 Rhipicephalus microplus ticks from Zebu cattle in Madagascar for Borrelia-specific DNA. Borrelia spp. DNA was detected in 21 Amblyomma variegatum ticks and 2 Rhipicephalus microplus ticks. One Borrelia found in one Rhipicephalus microplus showed close relationship to Borrelia theileri based on genetic distance and phylogenetic analyses on 16S rRNA and flaB sequences. The borreliae from Amblyomma variegatum could not be identified due to very low quantities of present DNA reflected by high cycle threshold values in real-time-PCR. It is uncertain whether these low numbers of Borrelia spp. are sufficient for transmission of infection from ticks to humans. In order to determine whether spirochaete infections are relevant in humans, blood samples of 1009 patients from the highlands of Madagascar with fever of unknown origin were screened for Borrelia spp. − and in addition for Leptospira spp. − by real-time PCR. No target DNA was detected, indicating a limited relevance of these pathogens for humans in the highlands of Madagascar. Highlights <UL> <LI> Borrelia spp. DNA was detected in Amblyomma (A.) variegatum ticks and Rhipicephalus (R.) microplus ticks in Madagascar. </LI> <LI> DNA of a Borrelia (B.) theileri-related borreliae was detected in R. microplus for the first time in Madagascar. </LI> <LI> Only low amounts of Borrelia spp. DNA were detected in the assessed ticks, indicating low numbers of organisms and indicating an at the most limited risk of transmission to humans. </LI> <LI> Borrelia spp. DNA was absent in the blood of fever patients from the highlands in Madagascar. </LI> <LI> Leptospira (L.) spp. DNA was absent in the blood of fever patients from the highlands in Madagascar. </LI> </UL>
3
Are brucellosis, Q fever and melioidosis potential causes of febrile illness in Madagascar?

Boone, Ides,Henning, Klaus,Hilbert, Angela,Neubauer, Heinrich,von Kalckreuth, Vera,Dekker, Denise Myriam,Schwarz, Norbert Georg,Pak, Gi Deok,Krü,ger, Andreas,Hagen, Ralf Matthias,Frickmann, Hagen Elsevier 2017 Acta tropica Vol.172 No.-
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Abstract Brucellosis, Q fever and melioidosis are zoonoses, which can lead to pyrexia. These diseases are often under-ascertained and underreported because of their unspecific clinical signs and symptoms, insufficient awareness by physicians and public health officers and limited diagnostic capabilities, especially in low-resource countries. Therefore, the presence of Brucella spp., Coxiella burnetii and Burkholderia pseudomallei was investigated in Malagasy patients exhibiting febrile illness. In addition, we analyzed zebu cattle and their ticks as potential reservoirs for Brucella and C. burnetii, respectively. Specific quantitative real-time PCR assays (qPCRs) were performed on 1020 blood samples drawn from febrile patients. In total, 15 samples (1.5%) were Brucella-positive, mainly originating from patients without travel history, while DNA from C. burnetii and Bu. pseudomallei was not detected. Anti-C. burnetii antibodies were found in four out of 201 zebu serum samples (2%), whereas anti-Brucella antibodies could not be detected. Brucella DNA was detected in a single zebu sample. Three out of 330 ticks analyzed (1%) were positively tested for C. burnetii DNA but with high Ct values in the qPCR assay. Our data suggest that zebus as well as Amblyomma and Boophilus ticks have to be considered as a natural reservoir or vector for C. burnetii, but the risk of cattle-to-human transmission is low. Since bovine brucellosis does not seem to contribute to human infections in Madagascar, other transmission routes have to be assumed.

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