DNA barcoding and delimitation of critically endangered indigenous and introduced tilapias (pisces cichlidae) of Pangani catchment, Northern Tanzania.

The Pangani catchment of Northern Tanzania harbours the critically endangered endemic tilapias of the genus Oreochromis. The introduction of non-native congenerics and consequent hybridization complicates taxa identification and phylogeny based on morphological systematics. We therefore morphologically and molecularly identified these tilapias and delimited their Molecular Operational Taxonomic Units (MOTUs) based on Cytochrome Oxidase Subunit I (CO1) gene for future management and conservation. A total of 132 indigenous and introduced tilapia specimens were morphologically identified, barcoded using the CO1 gene and delimited by Kimura 2 Parameter distance approaches, Automatic Barcode Gap Discovery (ABGD), Neighbour Joining (NJ) tree and haplotype analysis. Theoverall mean conspecific, congeneric and confamillial genetic distances based on the K2P model were 0.54%, 5.32% and 13.29% respectively. All taxa had a mean K2P distance < 2% and 90% (n = 10), were clearly delimited by the ABGD method. The NJ tree delimited tilapia taxa commensurate to the genetic distances depicted by DNA barcoding. However, DNA barcoding and NJ tree coherently failed to discriminate the morphologically distinct allopatric Oreochromis jipe and Oreochromis hunteri taxa. Moreover, the two methods depicted lack of monophyly in Oreochromis korogwe MOTUs implying that the taxon could consist of at least one MOTU. We conclude that the integration of morphological-based taxonomy and DNA barcoding among ichthyofaunal taxa herein will be invaluable in conservation and management of native tilapias in Pangani basin.

Detection of Porcine Cysticercosis in Meat Juice Samples from Infected Pigs.

BACKGROUND: Seroprevalence of porcine cysticercosis has been generally studied using enzyme-linked immunosorbent assays (ELISA) detecting either antigens or antibodies in sera. However, serum is not always readily available. The objective of this study was to assess the diagnostic potential of meat juice in detecting porcine cysticercosis using a cysticercosis antibody ELISA. METHODS: Sera and meat juice samples from 13 different organs/tissues were collected from nine pigs naturally infected with cysticercosis and from six uninfected pigs reared under hygienic conditions. The sensitivity of the cysticercosis antibody ELISA in detecting porcine cysticercosis in meat juice samples was compared to that in serum samples from the same pigs. RESULTS: Using sera, cysticercosis was detected in all nine pigs harbouring cysticerci, but not in those reared under hygienic conditions. The sensitivity of the ELISA was highest in meat juice extracted from the diaphragm (100%), heart (89%) and neck muscle (78%) of the nine infected pigs, whereas it varied between 0 and 44% in the other samples. CONCLUSION: To the best of our knowledge, this is the first study for T. solium cysticercosis serology to use meat juice. Our results show that meat juice from pig carcass organs or muscles is a promising diagnostic specimen for the detection of porcine cysticercosis. More studies including a large sample size of pigs with varying degrees of cysticercosis infection are needed to further prove this concept.

Soil contamination by Taenia solium egg DNA in rural villages in Kongwa district, Tanzania.

The presence ofTaenia solium DNA from eggs in soils around the households in four Tanzanian villages in Kongwa district were analysed in relation to seasonal fluctuations and infection risk implications. A total of 192 pooled soil samples from five sampling points per household were examined by droplet digital Polymerase Chain Reaction (ddPCR) from 96 pig-keeping households both during the dry and rainy seasons. The pooled samples were first processed by a flotation-double sieving technique, followed by screening for worm DNA employing universal primers targeting the mitochondrial cytochrome c oxidase subunit I (cox1) gene of human taeniid species and some other helminths. All DNA positive samples were later confirmed by a specific ddPCR probe assay targeting the mitochondrial cox1 gene of T. solium. A total of 17.2% (n = 33) samples were positive with the universal ddPCR, whereas T. solium DNA was confirmed by the specific ddPCR only in 3.1% (n = 3) of the surveyed households. The detection of T. solium DNA in this study spells out a low risk of exposure to T. solium eggs from contaminated household soil. Based on our results, ddPCR seems to be a promising technology for screening T. solium eggs in soil.

Validation of droplet digital Polymerase Chain Reaction for the detection and absolute quantification of Taenia solium eggs in spiked soil samples.

To enable the detection of taeniid eggs in environmental samples, a sensitive technology is required. In this study, we validated the effectiveness of a digital droplet Polymerase Chain Reaction (ddPCR) assay for detection, identification and absolute quantification of taeniid DNA from artificially contaminated soils with varying numbers of taeniid eggs using a set of universal primers, JB3 & JB4.5. The results showed that the number of cox1 copies detected increased gradually for both species with the number of taeniid eggs added to the different soil types. The highest cox1 DNA copies recovery for Taenia solium and T. lynciscapreoli was from the sand soil with lowest recovery being observed in clay soils. Therefore, ddPCR is a promising technology for screening of taeniid eggs from soil samples collected in the environment irrespective of the soil type and the number of eggs. The potential of the ddPCR protocol to detect taeniid egg DNA in spiked soil samples has great practical application for taeniid egg screening in soils from endemic areas. However, when universal primers are used in screening environmental samples, the identity of ddPCR positive samples must be confirmed by sequencing. In addition, more validation studies using species-specific primers and field soil samples is recommended.