Browsing by Author "Gwakisa, Paul"

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16S rRNA Amplicons Survey Revealed Unprecedented Bacterial Community in Solid Biomedical Wastes
(Science and Education Publishing, 2015) Mwaikono, Kilaza Samson; Maina, Solomon; Sebastian, Aswathy; Kapur, Vivek; Gwakisa, Paul
Despite known risks of inappropriate disposal of biomedical solid waste; most cities in developing countries are still disposing unsorted and untreated solid biomedical waste in common dumpsites. While many studies reported the presence of pathogens in fresh biomedical waste from hospitals, none has reported on the abundance and diversity of bacterial community in aged solid biomedical waste from a common dumpsite. A qualitative survey was done to identify types of solid biomedical waste on the dumpsite. Soils, sludge or washings of biomedical wastes were sampled. Total DNA was extracted and v4 region of 16S rRNA amplicons were sequenced using an Illumina MiSeq platform. A total of 1,706,442 sequences from 15 samples passed quality control. The number of sequences per sample ranged from 70664 to 174456 (mean 121765, SD 35853). Diversity was high with an InvSimpson index of 63 (Range 5 – 496, SD 121). Thirty five phyla were identified, but only 9 accounted for 96% of all sequences. The dominant phyla were Proteobacteria 37.4%, Firmicutes 34.4%, Bacteroidetes 14.1 %, Actinobacteria 5.6% and Chloroflex 1.7%. Catchall analysis predicted a mean of 9399 species per sample. Overall, 31402 operational taxonomic units (OTUs) were detected, however, only 19.8% (6,202) OTUs were found more than ten times. The most predominant OTUs were Proteinclasticum (10.4%), Acinetobacter (6.9), Halomonas (3.9), Pseudomonas (1.7%), Escherichia/Shigella 1.5% and Planococcus (1.3%). Proteiniclasticum spp and Acinetobacter spp were found in 67% (10/15) of all samples at relative abundance of 1%. Taxonomic-to-phenotype mapping revealed the presence of 36.2% related to bacteria involved in dehalogenation, 11.6% degraders of aromatic hydrocarbons, 14.8% chitin degraders, 8.5% chlorophenol degradation and Atrazine metabolism 8.3%. Taxonomy-to human pathogen mapping found 34% related to human pathogens and 39.4% were unknown. Conclusions There’s rich and diverse bacterial community in aged solid biomedical waste. Some of the predominant OTUs are related to bacteria of industrial use.We found a good number of OTUs mapping to human pathogens. Most of OTUs mapped to unknown metabolism and also to group unknown whether they human pathogens or not. To our knowledge, this is the first reports on bacteria related to industrial use from solid biomedical waste. This finding will facilitate to design further research using functional metagenomics to better understand the potential of bacteria from aged solid biomedical waste.
Application of magnetic cytosmear for the estimation of Plasmodium falciparum gametocyte density and detection of asexual stages in asymptomatic children
(Malaria Journal, 2016-02-24) Sumari, Deborah; Grimberg, Brian T.; Blankenship, D’Arbra; Mugasa, Joseph; Mugittu, Kefas; Moore, Lee; Gwakisa, Paul; Zborowski, Maciej
Background: Conventional malaria parasite detection methods, such as rapid diagnostic tests (RDT) and light microscopy (LM), are not sensitive enough to detect low level parasites and identification of gametocytes in the peripheral blood. A modified and sensitive laboratory prototype, Magnetic Deposition Microscopy (MDM) was developed to increase the detection of sub-microscopic parasitaemia and estimation of gametocytes density in asymptomatic school children. Methods: Blood samples were collected from 303 asymptomatic school children from seven villages in Bagamoyo district in Tanzania. Participants were screened for presence of malaria parasites in the field using RDT and MDM whereas further examination of malaria parasites was done in the laboratory by LM. LM and MDM readings were used to calculate densities and estimate prevalence of asexual and sexual stages of the parasite. Results: Plasmodium falciparum parasites (asexual and sexual stages) were detected in 23 (7.6 %), 52 (17.2 %), and 59 (19.5 %) out of 303 samples by LM, RDT and MDM respectively. Gametocytes were detected in 4 (1.3 %) and 12 (4.0 %) out of the same numbers of samples by LM, and MDM, respectively. Likewise, in vitro results conducted on two laboratory strains of P. falciparum, 3D7 and NF54 to assess MDM sensitivity on gametocytes detection and its application on concentrating gametocytes indicated that gametocytes were enriched by MDM by 10-fold higher than LM. Late stages of the parasite strains, 3D7 and NF54 were enriched by MDM by a factor of 20.5 and 35.6, respectively. MDM was more specific than LM and RDT by 87.5 % (95 %, CI 71.2–89.6 %) and 89.0 % (95 % CI 82.9–91.4) respectively. It was also found that MDM sensitivity was 62.5 % (95 % CI 49.5–71.8) when compared with RDT while with LM was 36.5 % (95 % CI 32.2–60.5). Conclusions: These findings provide strong evidence that MDM enhanced detection of sub-microscopic P. falciparum infections and estimation of gametocyte density compared to current malaria diagnostic tools. In addition, MDM is superior to LM in detecting sub-microscopic gametocytaemia. Therefore, MDM is a potential tool for low-level parasitaemia identification and quantification with possible application in malaria transmission research.
Assessing Risk Factors for Trypanosome Infections in Cattle in Wildlife Interface Areas in Northern Tanzania
(Journal of Infectious Diseases and Epidemiology, 2019-05-04) Ngongolo, Kelvin; Estes, Anna; Hudson, Peter; Gwakisa, Paul
Trypanosomosis is a vector-borne, tropical disease that causes mortality and morbidity in livestock and humans. In this study we investigated the risk factors for trypanosome infection in cattle in the Maasai Steppe of northern Tanzania. We assessed the influence of age, sex, herd size and history of treatment against trypanosomosis as risk factors of trypanosome infection. Cattle blood samples were collected from 150 cattle in three villages in the vicinity of Tarangire National Park, which acts as a reservoir of tsetse flies, the trypanosome vector. Parasite species were identified using a nested Polymerase Chain Reaction (n-PCR). Smaller herd sizes, young age (1-2 years), and male sex significantly increased the risk of trypanosome infections. Efforts to control trypanosome infection should be strategically based on location and season while considering age, treatment and herd size as risk factors.
CD5+ B lymphocytes are the main source of antibodies reactive with non-parasite antigens in Trypanosoma congolense-infected cattle
(Blackwell Science Ltd, 1997) Buza, Joram; Sileghem, Maarten; Gwakisa, Paul; Naessens, Jan
Mice infected with African trypanosomes produce exceptionally large amounts of serum IgM, a major part of which binds to non-trypanosome antigens such as trinitrophenol and single-strand DNA. In this paper, we describe that in cattle infected with Trypanosoma congolense and T. vivax, similar antibodies are found, although they bind mainly to protein antigens, such as b-galactosidase, ovalbumin and ferritin. The parasite non-specific IgM antibodies appear around the same time as the parasite-specific antibodies, but their origin and function are not clear.We tested the hypothesis that CD5+ B cells (or B-1 cells), which increase during trypanosome infections in cattle, are responsible for production of antibodies to non-trypanosome antigens. Splenic CD5+ and CD5− B cells from infected cattle were sorted and tested in a single cell blot assay. The numbers of immunoglobulin-secreting cells were similar in both B-cell populations. However, antibodies with reactivity for non-trypanosome antigens were significantly more prevalent in the CD5+ B-cell fraction and were exclusively IgM. The preference for production of these antibodies by CD5+ B cells and the expansion of this subpopulation during infections in cattle, strongly suggest that CD5+ B cells are the main source of trypanosome non-specific antibodies. We propose that these antibodies are natural, polyreactive antibodies that are predominantly secreted by CD5+ B cells. Since B-1 cells are up-regulated in many states of immune insufficiency, the immunosuppression associated with trypanosome infections may be responsible for the increase of this subset and the concomitant increase in trypanosome non-specific antibodies.
Detection of carrier state and genetic diversity of Theileria parva in ECF-vaccinated and naturally exposed cattle in Tanzania
(Elsevier B.V., 2019-08) Magulu, Emelesiana Cyprian; Kindoro, Fatuma; Mwega, Elisa; Kimera, Sharadhuli; Shirima, Gabriel; Gwakisa, Paul
Infection and Treatment Method (ITM) has been practiced in Tanzania for over 20 years as a prevention measure against East Coast Fever disease. It is known that ITM, like natural ECF infection, leads to a carrier state, whereby vaccinated cattle become asymptomatic carriers of the parasite. It is expected that ECF vaccination using ITM also leads to generation of combinations of vaccine specific Theileria parva and local strains that circulate in the field what contributes to an unknown level of parasite diversity. Moreover, the long term impact of ITM on carrier state and parasite diversity in cattle are largely unknown. To address this question blood was collected from ECF-vaccinated (n = 239) and unvaccinated (n = 97) cattle from Loiborsoit, Emboreet, Esilalei, Manyara ranch and Mswakini villages in the Maasai steppe of northern Tanzania, as well as Mruazi and Leila farms in Tanga in eastern Tanzania. Screening for T. parva using nested PCR revealed an overall prevalence of T. parva to be 34.5%, with a significant higher prevalence among ECF-vaccinated cattle. Using three VNTR markers (ms2, ms5 and MS7) higher parasite genetic diversity in terms of higher number of alleles and expected heterozygosity was shown in vaccinated than unvaccinated cattle. These parameters were highest in cattle from Manyara ranch. Nevertheless, the principle component analysis (PCoA) showed no distinct clustering patterns as most T. parva alleles clustered together throughout the four quadrants implying parasite homogeneity among the sampled populations. However, some of the parasite alleles closely clustered with Muguga vaccine alleles in two of the quadrants, consistent with closer genetic relatedness between the vaccine strains and the T. parva populations from the Maasai steppe. Likewise analysis of molecular variance (AMOVA) revealed most of the genetic variation (93%) being contained within populations with only 7% being among populations. This study therefore confirms the role of ECF vaccination in enhancing carrier state and T. parva diversity in vaccinated cattle populations. Higher T. parva diversity may play an important role in carrier cattle by way of restricting breakthrough infections from field parasite strains.
Identification of Bacillus anthracis, Brucella spp., and Coxiella burnetii DNA signatures from bushmeat
(Springer Nature Limited, 2021-07-21) Katani, Robab; Schilling, Megan; Lyimo, Beatus; Eblate, Ernest; Martin, Andimile; Tonui, Triza; Cattadori, Isabella; Francesconi, Stephen; Estes, Anna; Rentsch, Dennis; Srinivasan, Sreenidhi; Lyimo, Samson; Munuo, Lidia; Tiambo, Christian; Stomeo, Francesca; Gwakisa, Paul; Mosha, Fausta; Hudson, Peter; Buza, Joram; Kapur, Vivek
Meat from wildlife species (bushmeat) represents a major source of dietary protein in low- and middle-income countries where humans and wildlife live in close proximity. Despite the occurrence of zoonotic pathogens in wildlife, their prevalence in bushmeat remains unknown. To assess the risk of exposure to major pathogens in bushmeat, a total of 3784 samples, both fresh and processed, were collected from three major regions in Tanzania during both rainy and dry seasons, and were screened by real-time PCR for the presence of DNA signatures of Bacillus anthracis (B. anthracis), Brucella spp. (Brucella) and Coxiella burnetii (Coxiella). The analysis identified DNA signatures of B. anthracis (0.48%), Brucella (0.9%), and Coxiella (0.66%) in a total of 77 samples. Highest prevalence rates of B. anthracis, Brucella, and Coxiella were observed in wildebeest (56%), dik-dik (50%), and impala (24%), respectively. Fresh samples, those collected during the rainy season, and samples from Selous or Serengeti had a greater relative risk of being positive. Microbiome characterization identified Firmicutes and Proteobacteria as the most abundant phyla. The results highlight and define potential risks of exposure to endemic wildlife diseases from bushmeat and the need for future investigations to address the public health and emerging infectious disease risks associated with bushmeat harvesting, trade, and consumption.
iMAP: an integrated bioinformatics and visualization pipeline for microbiome data analysis
(BMC Bioinformatics, 2019) Buza, Teresia; Tonui, Triza; Stomeo, Francesca; Tiambo, Christian; Katani, Robab; Schilling, Megan; Lyimo, Beatus; Gwakisa, Paul; Cattadori, Isabella; Buza, Teresia; Kapur, Vivek
Background: One of the major challenges facing investigators in the microbiome field is turning large numbers of reads generated by next-generation sequencing (NGS) platforms into biological knowledge. Effective analytical workflows that guarantee reproducibility, repeatability, and result provenance are essential requirements of modern microbiome research. For nearly a decade, several state-of-the-art bioinformatics tools have been developed for understanding microbial communities living in a given sample. However, most of these tools are built with many functions that require an in-depth understanding of their implementation and the choice of additional tools for visualizing the final output. Furthermore, microbiome analysis can be time-consuming and may even require more advanced programming skills which some investigators may be lacking. Results: We have developed a wrapper named iMAP (Integrated Microbiome Analysis Pipeline) to provide the microbiome research community with a user-friendly and portable tool that integrates bioinformatics analysis and data visualization. The iMAP tool wraps functionalities for metadata profiling, quality control of reads, sequence processing and classification, and diversity analysis of operational taxonomic units. This pipeline is also capable of generating web-based progress reports for enhancing an approach referred to as review-as-you-go (RAYG). For the most part, the profiling of microbial community is done using functionalities implemented in Mothur or QIIME2 platform. Also, it uses different R packages for graphics and R-markdown for generating progress reports. We have used a case study to demonstrate the application of the iMAP pipeline. Conclusions: The iMAP pipeline integrates several functionalities for better identification of microbial communities present in a given sample. The pipeline performs in-depth quality control that guarantees high-quality results and accurate conclusions. The vibrant visuals produced by the pipeline facilitate a better understanding of the complex and multidimensional microbiome data. The integrated RAYG approach enables the generation of web-based reports, which provides the investigators with the intermediate output that can be reviewed progressively. The intensively analyzed case study set a model for microbiome data analysis.
iMAP: an integrated bioinformatics and visualization pipeline for microbiome data analysis
(BioMed Central, 2019-07-03) Buza, Teresia; Tonui, Triza; Stomeo, Francesca; Tiambo, Christian; Katani, Robab; Schilling, Megan; Lyimo, Beatus; Gwakisa, Paul; Cattadori, Isabella; Buza, Joram; Kapur, Vivek
One of the major challenges facing investigators in the microbiome field is turning large numbers of reads generated by next-generation sequencing (NGS) platforms into biological knowledge. Effective analytical workflows that guarantee reproducibility, repeatability, and result provenance are essential requirements of modern microbiome research. For nearly a decade, several state-of-the-art bioinformatics tools have been developed for understanding microbial communities living in a given sample. However, most of these tools are built with many functions that require an in-depth understanding of their implementation and the choice of additional tools for visualizing the final output. Furthermore, microbiome analysis can be time-consuming and may even require more advanced programming skills which some investigators may be lacking. Results We have developed a wrapper named iMAP (Integrated Microbiome Analysis Pipeline) to provide the microbiome research community with a user-friendly and portable tool that integrates bioinformatics analysis and data visualization. The iMAP tool wraps functionalities for metadata profiling, quality control of reads, sequence processing and classification, and diversity analysis of operational taxonomic units. This pipeline is also capable of generating web-based progress reports for enhancing an approach referred to as review-as-you-go (RAYG). For the most part, the profiling of microbial community is done using functionalities implemented in Mothur or QIIME2 platform. Also, it uses different R packages for graphics and R-markdown for generating progress reports. We have used a case study to demonstrate the application of the iMAP pipeline. Conclusions The iMAP pipeline integrates several functionalities for better identification of microbial communities present in a given sample. The pipeline performs in-depth quality control that guarantees high-quality results and accurate conclusions. The vibrant visuals produced by the pipeline facilitate a better understanding of the complex and multidimensional microbiome data. The integrated RAYG approach enables the generation of web-based reports, which provides the investigators with the intermediate output that can be reviewed progressively. The intensively analyzed case study set a model for microbiome data analysis.
Influence of Land Cover and Host Species on Trypanosome Infection in Cattle and its Socio-Economic Impacts to Pastoralists of the Maasai Steppe, Tanzania
(Journal of Infectious Diseases and Epidemiology, 2020-01-27) Ngongolo, Kelvin; Shirima, Gabriel; Mpolya, Emmanuel; Estes, Anna; Hudson, Peter; Gwakisa, Paul
Introduction Trypanosome infections result into trypanosomosis in cattle and this is an infection detrimental to pastoralist income. The patterns of transmission are thought to be influenced by ecological factors including wildlife and land cover. We assessed the influence of the relative abundance of wildlife and land cover (cultivation and habitat type) on the presence of trypanosome infections in replicated cattle herds of the Maasai Steppe. Methodology We undertook a cohort field study in three villages of the Maasai Steppe: Sukuro, Kimotorok and Oltukai. The study took place in July 2017 and October 2017 and utilized 50 cattle from each village. Pastoralists were asked questions during each visit when blood sampled were taken to seek their viewpoint on the relative abundance of wildlife, habitat types and cultivation observed in the areas their cattle grazed. In addition, the percentage cover of cultivated land and habitat types in the grazing areas were determined during field visits and participatory mapping with pastoralists. A systematic review was used to understand the socio-economic importance of trypanosomosis. The species of trypanosomes in cattle were identified using nested Polymerase chain reaction (n-PCR). Results There was a significant and positive association between the presence of trypanosome infection and the abundance of wildlife within grazing areas, in particular the abundance of buffaloes (Odd Ratio > 1, P = 0.038, 95% CI 1.26 to 1.38) when cattle grazed in woodland habitats. Cultivation on grazing areas had a negative association with the presence of trypanosome infections (R < 1, P = 0.001, 95% CI 0.0614 to 0.0986) in cattle but this varied between villages. A systematic review showed that trypanosomosis had socio-economic impacts such as loss of income, reduced quality, and quantity of livestock products, management cost, and inadequate provisions of socio-services and potential zoonotic transmission to humans. Conclusion & recommendations The socio-economic impacts of trypanosomosis will continue to be a challenge to pastoralists when cattle are grazed close to wildlife areas which are infested with tsetse fly habitats. Control strategies for trypanosome infection in cattle on the Maasai Steppe should consider the interaction of cattle with ecological factors.
Molecular species identification of bushmeat recovered from the Serengeti ecosystem in Tanzania.
(PLOS ONE, 2020-09-14) Schilling, Megan; Estes, Anna; Eblate, Ernest; Martin, Andimile; Rentsch, Dennis; Katani, Robab; Joseph, Asteria; Kindoro, Fatuma; Lyimo, Beatus; Radzio-Basu, Jessica; Cattadori, Isabella; Hudson, Peter; Kapur, Vivek; Gwakisa, Paul; Buza, Joram
Bushmeat harvesting and consumption represents a potential risk for the spillover of endemic zoonotic pathogens, yet remains a common practice in many parts of the world. Given that the harvesting and selling of bushmeat is illegal in Tanzania and other parts of Africa, the supply chain is informal and may include hunters, whole-sellers, retailers, and individual resellers who typically sell bushmeat in small pieces. These pieces are often further processed, obscuring species-identifying morphological characteristics, contributing to incomplete or mistaken knowledge of species of origin and potentially confounding assessments of pathogen spillover risk and bushmeat offtake. The current investigation sought to identify the species of origin and assess the concordance between seller-reported and laboratory-confirmed species of origin of bushmeat harvested from in and around the Serengeti National Park in Tanzania. After obtaining necessary permits, the species of origin of a total of 151 bushmeat samples purchased from known intermediaries from 2016 to 2018 were characterized by PCR and sequence analysis of the cytochrome B (CytB) gene. Based on these sequence analyses, 30%, 95% Confidence Interval (CI: 24.4-38.6) of bushmeat samples were misidentified by sellers. Misreporting amongst the top five source species (wildebeest, buffalo, impala, zebra, and giraffe) ranged from 20% (CI: 11.4-33.2) for samples reported as wildebeest to 47% (CI: 22.2-72.7) for samples reported as zebra although there was no systematic bias in reporting. Our findings suggest that while misreporting errors are unlikely to confound wildlife offtake estimates for bushmeat consumption within the Serengeti ecosystem, the role of misreporting bias on the risk of spillover events of endemic zoonotic infections from bushmeat requires further investigation.
Prevalence and Antimicrobial Resistance Phenotype of Enteric Bacteria from a Municipal Dumpsite
(Science and Education Publishing, 2015) Mwaikono, Kilaza Samson; Maina, Solomon; Gwakisa, Paul
The objective of the study was to determine the prevalence and antibiotic resistance phenotype of enteric bacteria from the municipal dumpsite. A qualitative survey of the dumpsite was conducted to identify types of solid wastes and nature of interaction on the dumpsite. Samples were collected from different type of solid waste, including domestic waste (Dom), solid biomedical waste (Biom), river sludge near the dumpsite (Riv) and faecal material of pigs scavenging on the dumpsite (FecD). A control sample was collected from faecal material of pigs initially reared indoor (FecI) and shifted to scavenging on the dumpsite (FecIF). Total genomic DNA was extracted, and the 16S rRNA gene was amplified, sequenced and used to study prevalence of enteric bacteria. The same sample was used to isolate enteric bacteria that were later tested to 8 different antibiotics for their susceptibility phenotype. Solid wastes are not sorted in Arusha municipal. There was high interaction between animals and humans on the dumpsite. A total of 219 enteric bacteria from 75 genera were identified. Escherichia sp and Shigella sp (12%), Bacillus sp (11%) and Proteiniclasticum (4%) were the predominant genera. Most of the Escherichia sp, Shigella sp and Bacillus were from FecD, while Proteiniclasticum spp was from Biom. Some isolates from FecD had 99% sequence similarity to pathogenic Escherichia furgosonii, Shigella sonnei, Enterococcus faecium and Escherichia coli O154:H4. Over 50% of the isolates were resistant to Penicillin G, Ceftazidime and Nalidixic Acid. Ciprofloxacin and Gentamycin were the most effective antibiotics with 81% and 79% susceptible isolates, respectively. Of all the isolates, 56% (45/80) were multidrug resistant. Escherichia sp and Bacillus sp (12 isolates each) constituted a large group of multidrug resistant bacteria. All Pseudomonas sp from Biom and FecD were multidrug resistant. There is high prevalence of antibiotic resistant enteric bacteria on the dumpsite. We report possible risks of spreading antibiotic resistant bacteria/genes from the dumpsite to clinical settings through animals and humans interacting on the dumpsite. This finding calls for a comprehensive research to study the shared resistome in bacteria from the environment, humans and animals using PCR and metagenomic based approaches to identify prevalence of known and capture new resistant genes.