Conjugation of Microcystins with Thiols Is Reversible: Base-Catalyzed Deconjugation for Chemical Analysis.

Microcystins are potent cyclic heptapeptide toxins found in many freshwater cyanobacteria. Most microcystins contain an α,ß-unsaturated amide that can react with thiol-containing amino acids, peptides, and proteins in vivo and in vitro. While soluble conjugates formed from small peptides can be extracted and analyzed directly by LC-MS, microcystins conjugated to proteins are analyzed after oxidative cleavage of their Adda side chains, but information on which microcystin analogues were present is lost. Observations during the development of thiol-derivatization-based LC-MS methods for microcystin analysis indicated that the reaction of thiols with microcystins was reversible. The kinetics of deconjugation was investigated with mercaptoethanol as a model thiol to identify suitable reaction conditions. A range of microcystins conjugated to mercaptoethanol, methanethiol, cysteine, and glutathione were then successfully deconjugated, demonstrating the feasibility of releasing conjugated forms of microcystins for chemical analysis. Reagents for removing the released thiols or for trapping the released microcystins increased the reaction rate. Optimization of methodologies based on this reaction should increase the method's utility for measuring free and conjugated microcystins. The results also indicate that thiol-conjugated microcystins slowly release free microcystins, even at neutral pH, with consequences for assessment of toxin exposure, metabolism, and trophic transfer. A range of other common natural and environmental toxins, such as deoxynivalenol and acrylamide, also contain α,ß-unsaturated carbonyl groups and can be expected to behave in a similar manner.

Epidemiology and RAPD-PCR typing of thermophilic campylobacters from children under five years and chickens in Morogoro Municipality, Tanzania.

BACKGROUND: Campylobacter species are gram negative and flagellated bacteria under the genus Campylobacter, family Campylobacteriaceae. These pathogens cause zoonotic infections among human and animal populations. This study was undertaken between December 2006 and May 2007 to determine prevalence, risk factors and genetic diversity of thermophilic Campylobacter isolates from children less than 5 years and chickens in Morogoro Municipality, Tanzania. METHODS: The Skirrow's protocol was used for isolation and identification of Campylobacter from 268 human stool specimens and 419 chicken cloacal swabs. Patient biodata and risk factors associated with human infection were also collected. Genetic diversity of Campylobacter isolates was determined by a RAPD-PCR technique using OPA 11 primer (5'-CAA TCG CCG T-3'). Phylogenetic analysis and band pattern comparison were done by Bionumerics software and visual inspection. RESULTS: Stool samples from 268 children and 419 cloacal swabs from chickens were analyzed. Prevalence of thermophilic Campylobacters in children was 19% with higher isolation frequency (p = 0.046) in males (23.5%) than females (13.8%). Campylobacter jejuni (78.4%) was more isolated (p = 0.000) than C. coli (19.6%) and 2% were unidentified isolates. In chickens, the prevalence was 42.5% with higher isolation rate (p = 0.000) of C. jejuni (87%) than C. coli (13%). Campylobacters were more frequently recovered (p = 0.000) from indigenous/ local chickens (75.0%) followed by cockerels (52.2%), broilers (50.0%) and lowest in layers (22.7%). Keeping chickens without other domestic animals concurrently (p = 0.000), chicken types (p = 0.000) and flock size (p = 0.007) were risk factors for infection in chickens. One hundred and fifty two (152) thermophillic Campylobacter isolates were genotyped by RAPD-PCR of which 114 were C. jejuni (74 from chickens and 40 humans) and 38 C. coli (28 from chickens and 10 humans). Comparison of Campylobacter isolates from children and chickens revealed high diversity with only 6.1% of C. jejuni and 5.3% of C. coli being 100% genetically similar. CONCLUSIONS: This study has recorded high prevalence of thermophilic Campylobacter in children less than 5 years and chickens in Morogoro municipality. The observed genetic similarity among few C. jejuni and C. coli isolates from children and chicken suggests existence of cross transmission of these pathogens between children under 5 years and chickens.

Semiquantitative Risk Evaluation Reveals Drivers of African Swine Fever Virus Transmission in Smallholder Pig Farms and Gaps in Biosecurity, Tanzania.

African swine fever (ASF) has remained persistent in Tanzania since the early 2000s. Between 2020 and 2021, pig farms in twelve districts in Tanzania were infected with ASF, and ≥4,804 pigs reportedly died directly due to the disease with disruption to livelihoods. We conducted semiquantitative field investigations and rapid risk assessment (RRA) to understand the risk factors and drivers of ASF virus (ASFV) amplification and transmission in smallholder pig farms, and determine the gaps in biosecurity through hazard profiling, focus group discussions and expert opinion. Outbreaks were connected by road and aligned along the pig product value chain and reported in the northern, central, and southern parts of Tanzania. The patterns of outbreaks and impacts differed among districts, but cases of ASF appeared to be self-limiting following significant mortality of pigs in farms. Movement of infected pigs, movement of contaminated pig products, and fomites associated with service providers, vehicles, and equipment, as well as the inadvertent risks associated with movements of animal health practitioners, visitors, and scavengers were the riskiest pathways to introduce ASFV into smallholder pig farms. Identified drivers and facilitators of risk of ASFV infection in smallholder pig farms were traders in whole pigs, middlemen, pig farmers, transporters, unauthorized animal health service providers, and traders in pork. All identified pig groups were susceptible to ASFV, particularly shared adult boars, pregnant and lactating sows, and other adult females. The risk of ASF for smallholder pig farms in Tanzania remains very high based on a systematic risk classification. The majority of the farms had poor biosecurity and no single farm implemented all identified biosecurity measures. Risky practices and breaches of biosecurity in the pig value chain in Tanzania are profit driven and are extremely difficult to change. Behavioural change communication must target identified drivers of infections, attitudes, and practices.

Occurrence and Risk Factors of Brucellosis in Commercial Cattle Farms from Selected Districts of the Eastern Coast Zone, Tanzania.

Brucellosis is a disease of major socio-economic importance worldwide, particularly in low-income countries. This retrospective study aimed to estimate seroprevalence and risk factors associated with brucellosis in commercial cattle farms in the eastern coast zone of Tanzania (ECZT). A total of 1,052 serum samples collected from 20 commercial farms were subjected to rose bengal plate test (RBPT) and indirect enzyme-linked immunosorbent assay (i-ELISA). Descriptive analysis was employed to determine frequencies and proportions. To establish risk factors, a multivariate logistic regression analysis was carried out using a backward elimination procedure, following a univariate analysis, with 0.1 set as a cut-off point for the selection of putative risk factors. Agreement between RBPT and i-ELISA was determined using a Kappa coefficient (κ). The overall animal-level seroprevalence was 25.9% based on i-ELISA. Logistic regression analysis revealed that odds of infection were significantly higher in females (OR = 1.8, 95% CI: 1.2-2.5, p = 0.002) and in young animals than in adults (OR = 3.6, CI: 2.1-6.2, p < 0.001). In addition, odds of infection were higher during the wet season (OR = 3.4, CI: 3.2-5.2, p < 0.001), in cattle reared in rural farms (OR = 4.8, CI: 2.0-11.5, p < 0.001), in cattle reared in areas, not in contact with wildlife (OR = 2.9, CI: 1.4-2.3, p = 0.004), and in medium-sized farms (OR = 12.5, CI: 6.9-22.9, p < 0.001). These findings confirm that bovine brucellosis was prevalent among commercial cattle farms in the ECZT, posing a serious public health concern to the community living in these settings. The one health approach should be adopted for effective control of brucellosis.

Thiol derivatization for LC-MS identification of microcystins in complex matrices.

Microcystins are a group of cyclic heptapeptides originating from cyanobacteria. Cyanobacteria also produce a range of peptides and other compounds that can result in complex chromatograms when samples are analyzed by LC-MS. Derivatization with appropriate thiols (e.g., mercaptoethanol) of the olefin in the α,ß-unsaturated amide present in most microcystins was shown to simplify analysis of LC-MS chromatograms of sample extracts, making it much easier to identify peaks corresponding to candidate microcystins. Furthermore, interpretation of MS(2) spectra was facilitated by addition of the mass associated with the thiol to the α,ß-unsaturated amide of microcystins. Cyanotoxins containing Mdha or Dha reacted readily with thiols, whereas Mser, Ser, Mdhb, and thiol-derivatives of Mdha or Dha did not react under the conditions used. This approach therefore provides a convenient LC-MS method to obtain evidence for the presence of Mdha or Dha and can likely be used to differentiate between the isobaric amino acids Mdha and Dhb in candidate cyanotoxin peaks. When O-(2-mercaptoethyl)-O'-methyl-hexa(ethylene glycol) (MEMHEG) (M(w)t. 356) was used as the thiol, the resulting derivatives eluted in an LC-MS mass window that was largely free of interferences. This approach simplifies detection of candidate microcystin analogues even in the presence of complex mixtures of coeluting components. The method was used for qualitative analysis of a Microcystis aeruginosa culture from Lake Naivasha, Kenya, and the results were verified using precursor-ion scanning and high-resolution mass spectrometry.

Antimicrobial Use, Residues, Resistance and Governance in the Food and Agriculture Sectors, Tanzania.

All infections are potentially curable as long as the etiological agents are susceptible to antimicrobials. The increased rate at which antimicrobials are becoming ineffective is a global health risk of increasing concern that threatens withdrawal of beneficial antimicrobials for disease control. The increased demand for food of animal origin, in particular eggs, meat and milk has led to intensification and commercial production systems where excessive use and misuse of antimicrobials may prevail. Antimicrobials, handled and used by farmers and animal attendants with no formal education, may be predisposed to incorrect dosages, misuse, incorrect applications and non-adherence to withdrawal periods. This study was conducted to assess the regulatory roles and governance of antimicrobials, establish the pattern and extent of their use, evaluate the antimicrobial residues and resistance in the food animals and crop agriculture value chains, and relate these findings to existing strategies in place for combating the emergence of antimicrobial resistance in Tanzania. A multimethod approach (desk review, field study and interviews) was used. Relevant establishments were also visited. High levels of resistance to penicillin G, chloramphenicol, streptomycin and oxytetracycline have been reported, especially for Actinobacter pyogenes, Staphylococcus hyicus, Staphylococcus intermedius and Staphylococcus aureus from dairy cattle with mastitis and in humans. Similar trends were found in poultry where eggs and meat are contaminated with Escherichia coli strains resistant to amoxicillin + clavulanate, sulphamethoxazole and neomycin. An increasing trend of emerging multidrug resistant E. coli, Klebsiella pneumoniae, Staphylococcus aureus and Salmonella was also found in food animals. An increase in methicillin resistant Staphlococcus aureus (MRSA) and extended-spectrum beta-lactamase (ESBL) in the livestock sector in Tanzania have been reported. The pathogens isolated in animals were resistant to ampicillin, augmentin, gentamicin, co-trimoxazole, tetracycline, amoxicillin, streptomycin, nalidixic acid, azithromycin, chloramphenicol, tylosin, erythromycin, cefuroxime, norfloxacin and ciprofloxacin. An increased usage of antimicrobials for prophylaxis, and therapeutics against pathogens and for growth promotion in livestock, aquaculture and crop production were observed. A One Health strategic approach is advocated to combat antimicrobial resistance (AMR) in the food and agriculture sectors in Tanzania. Practical recommendations include (a) legislation review and implementation; (b) antimicrobial use (AMU), AMR and antimicrobial residue (AR) awareness and advocacy among stakeholders along the value chain; (c) strengthening of surveillance and monitoring programs for AMU, AMR and AR; (d) enhanced development and use of rapid and innovative diagnostic tests and the promotion of biosecurity principles; and (e) good husbandry practices. The utilization of this information to improve public health policies and reduce the burden of AMR will be beneficial.

Multisectoral cost analysis of a human and livestock anthrax outbreak in Songwe Region, Tanzania (December 2018-January 2019), using a novel Outbreak Costing Tool.

OBJECTIVES: We applied a novel Outbreak Costing Tool (OCT), developed by the US Centers for Disease Control and Prevention (CDC), to estimate the costs of investigating and responding to an anthrax outbreak in Tanzania. We also evaluated the OCT's overall utility in its application to a multisectoral outbreak response. METHODS: We collected data on direct costs associated with a human and animal anthrax outbreak in Songwe Region (December 2018 to January 2019) using structured questionnaires from key-informants. We performed a cost analysis by entering direct costs data into the OCT, grouped into seven cost categories: labor, office, travel and transport, communication, laboratory support, medical countermeasures, and consultancies. RESULTS: The total cost for investigating and responding to this outbreak was estimated at 102,232 United States dollars (USD), with travel and transport identified as the highest cost category (62,536 USD) and communication and consultancies as the lowest, with no expenditure, for the combined human and animal health sectors. CONCLUSIONS: Multisectoral investigation and response may become complex due to coordination challenges, thus allowing escalation of public health impacts. A standardized framework for collecting and analysing cost data is vital to understanding the nature of outbreaks, in anticipatory planning, in outbreak investigation and in reducing time to intervention. Pre-emptive use of the OCT will also reduce overall and specific (response period) intervention costs for the disease. Additional aggregation of the costs by government ministries, departments and tiers will improve the use of the tool to enhance sectoral budget planning for disease outbreaks in a multisectoral response.

Reservoir dynamics of rabies in south-east Tanzania and the roles of cross-species transmission and domestic dog vaccination.

Understanding the role of different species in the transmission of multi-host pathogens, such as rabies virus, is vital for effective control strategies. Across most of sub-Saharan Africa domestic dogs Canis familiaris are considered the reservoir for rabies, but the role of wildlife has been long debated. Here we explore the multi-host transmission dynamics of rabies across south-east Tanzania.Between January 2011 and July 2019, data on probable rabies cases were collected in the regions of Lindi and Mtwara. Hospital records of animal-bite patients presenting to healthcare facilities were used as sentinels for animal contact tracing. The timing, location and species of probable rabid animals were used to reconstruct transmission trees to infer who infected whom and the relative frequencies of within- and between-species transmission.During the study, 688 probable human rabies exposures were identified, resulting in 47 deaths. Of these exposures, 389 were from domestic dogs (56.5%) and 262 from jackals (38.1%). Over the same period, 549 probable animal rabies cases were traced: 303 in domestic dogs (55.2%) and 221 in jackals (40.3%), with the remainder in domestic cats and other wildlife species.Although dog-to-dog transmission was most commonly inferred (40.5% of transmission events), a third of inferred events involved wildlife-to-wildlife transmission (32.6%), and evidence suggested some sustained transmission chains within jackal populations.A steady decline in probable rabies cases in both humans and animals coincided with the implementation of widespread domestic dog vaccination during the first 6 years of the study. Following the lapse of this program, dog rabies cases began to increase in one of the northernmost districts. Synthesis and applications. In south-east Tanzania, despite a relatively high incidence of rabies in wildlife and evidence of wildlife-to-wildlife transmission, domestic dogs remain essential to the reservoir of infection. Continued dog vaccination alongside improved surveillance would allow a fuller understanding of the role of wildlife in maintaining transmission in this area. Nonetheless, dog vaccination clearly suppressed rabies in both domestic dog and wildlife populations, reducing both public health and conservation risks and, if sustained, has potential to eliminate rabies from this region.

Occurrences of thermophilic Campylobacter in cattle slaughtered at Morogoro municipal abattoir, Tanzania.

An investigation was conducted in Morogoro municipality to assess the likelihood of slaughter cattle posing public health risk of contaminating carcasses with thermophilic Campylobacter. Butchers and meat shopkeepers were interviewed on source of slaughter cattle, method of animal and carcass transportation, carcass dressing, meat storage facilities, access to clean water and availability of food hygiene practices. Faecal samples were collected from 107 slaughter cattle and after slaughter; four different parts of dressed carcasses (i.e. from ham, neck, pelvis and thigh muscles) were also sampled. In addition 107 cattle meat samples for Campylobacter culture were collected in different meat shops. The samples were subjected to standard bacteriological examination using Skirrows protocol. It was found that cattle slaughter, dressing and meat handling in meatshops was done under unhygienic condition. Thermophilic Campylobacter prevalence in slaughter cattle was 5.6% while contamination rate of dressed carcasses and cattle meat at shops was 9.3% and 1.9%, respectively. The majority of thermophilic Campylobacter isolated were C. jejuni (88.9%) while C. coli was isolated at 11.1%. Findings of this study suggest possibility of humans acquiring zoonotic Campylobacter infections from cattle meat particularly when meat preparation and processing is not done properly. More work is required to establish the magnitude of zoonotic enteric Campylobacteriosis in humans and epidemiological role of cattle and other animals in the study area.

Where and when to vaccinate? Interdisciplinary design and evaluation of the 2018 Tanzanian anti-rabies campaign.

OBJECTIVES: Hoping to improve health-related effectiveness, a two-phase vaccination against rabies was designed and executed in northern Tanzania in 2018, which included geo-epidemiological and economic perspectives. METHODS: Considering the local bio-geography and attempting to rapidly establish a protective ring around a city at risk, the first phase intervened on sites surrounding that city, where the population density was lower than in the city at risk. The second phase vaccinated a rural area. RESULTS: No rabies-related case has been reported in the vaccinated areas for over a year post-immunisation; hence, the campaign is viewed as highly cost-effective. Other metrics included: rapid implementation (concluded in half the time spent on other campaigns) and the estimated cost per protected life, which was 3.28 times lower than in similar vaccinations. CONCLUSIONS: The adopted design emphasised local bio-geographical dynamics: it prevented the occurrence of an epidemic in a city with a higher demographic density than its surrounding area and it also achieved greater effectiveness than average interventions. These interdisciplinary, policy-oriented experiences have broad and immediate applications in settings of limited and/or time-sensitive (expertise, personnel, and time available to intervene) resources and conditions.

Ecological niche modeling as a tool for prediction of the potential geographic distribution of Bacillus anthracis spores in Tanzania.

INTRODUCTION: Anthrax is caused by the spore-forming, Gram-positive bacterium Bacillus anthracis. The aim of this study was to predict the potential distribution of B. anthracis in Tanzania and produce epidemiological evidence for the management of anthrax outbreaks in the country. METHODS: The Maxent algorithm was used to predict areas at risk of anthrax outbreaks based on the occurrence and environmental data in Arusha and Kilimanjaro regions; the model was later transferred to predict the entire country. Seventy percent of the occurrence data were used to train the model, while 30% were used for model evaluation. RESULTS: Four regions of northern Tanzania are predicted to have a high risk for anthrax outbreaks, while the southern and western regions had low-risk areas. Soil type (56.5%), soil pH (23.7%), and isothermally (10.4%) were the most important variables for the model prediction, and the most significant soil types were solonetz, fluvisols, and lithosols. CONCLUSIONS: A strong risk level across districts of the Tanzania mainland was identified in this study. A total of 18 districts in Tanzania Mainland are predicted to be at very high risk of an anthrax outbreak occurrence. These findings are important for policymakers to effectively mount targeted control measures for anthrax outbreaks in Tanzania.

Risk factors for human cutaneous anthrax outbreaks in the hotspot districts of Northern Tanzania: an unmatched case-control study.

Bacillus anthracis is an aerobic, Gram-positive and spore-forming bacterium, which causes anthrax in herbivores. Humans get infected after coming into contact with infected animals' products. An unmatched case-control study was conducted to identify the importance of demographic, biological and/or behavioural factors associated with human cutaneous anthrax outbreaks in the hotspot areas of Northern Tanzania. A semi-structured questionnaire was administered to both cases and controls. The age range of participants was 1-80 years with a median age of 32 years. In the younger group (1-20 years), the odds of being infected were 25 times higher in the exposed group compared to the unexposed group (OR= 25, 95% CI = 1.5-410). By contrast, the odds of exposure in the old group (≥20 years) were three times lower in the exposed group compared to the unexposed group (OR = 3.2, 95% CI = 1.28-8.00). Demographic characteristics, sleeping on animal's skins, contacting with infected carcasses through skinning and butchering, and not having formal education were linked to exposure for anthrax infection. Hence, a One Health approach is inevitable for the prevention and control of anthrax outbreaks in the hotspot areas of Northern Tanzania.

Antibiotic susceptibilities of indicator bacteria Escherichia coli and Enterococci spp. isolated from ducks in Morogoro Municipality, Tanzania.

OBJECTIVE: To estimate the prevalence of antibiotic resistance in indicator bacteria Escherichia coli and Enterococci isolated from duck faeces in Morogoro Municipality, Tanzania. RESULTS: Escherichia coli and Enterococcus isolation rates from ducks faeces were 91 and 100% respectively. The prevalence of antibiotic resistance of E. coli and Enterococcus was 70.3 and 42%, respectively. E. coli resistant to four antibiotics were 28 (30.8%) and showed high resistance to ampicillin (81.3), tetracycline (75.8) and trimethoprim-sulphamethoxine (62.3). Multiple antibiotic resistance of Enterococcus were more than 65%. High resistance rates shown by Enterococcus were observed in rifampin (62%), ampicillin (62%) and tetracycline (42%). Almost all farmers (92.3%) left their ducks to scavenge for food around their houses. Antibiotics used in animal treatments were oxytetracyclines, sulfonamides, penicillin dihydrostreptomycin while in humans were tetracycline, ampicillin, and amoxicillin.

A survey of the causes of cattle organs and/or carcass condemnation, financial losses and magnitude of foetal wastage at an abattoir in Dodoma, Tanzania.

Slaughterhouses provide a safeguard that prevents the public from consuming meat of poor quality or meat which may be infected with zoonotic diseases. This work reviews a 3-year database of cattle that were slaughtered and inspected between 2010 and 2012 at Dodoma abattoir, Tanzania. In addition, meat inspection was undertaken for 1 month (December 2013). The aim of this study was to establish causes of organ and carcass condemnations and their financial implications as well as the magnitude of slaughter of pregnant cows at Dodoma abattoir. During retrospective study, it was found that a total of 9015 (10.5%) lungs, 6276 (7.3%) intestines, 5402 (6.3%) livers, 3291 (3.8%) kidneys and 41 (0.05%) carcasses were condemned. Pulmonary emphysema (3.4%), fasciolosis (4.5%), pimply gut (5.7%), kidney congenital cysts (1.9%) and hydatidosis (3.1%) were major causes of organ condemnations. This large number of condemned edible organs and/or carcasses implies that public health considerations result in deprivation of valuable protein. Occurrence of hydatidosis, cysticercosis, fasciolosis and tuberculosis illustrates the possible public health problem and presence of environmental infections. Of the 794 cows slaughtered in December 2013, 46% were pregnant. Financial loss as a result of organ and/or carcass condemnations was estimated at $9892. Condemnation of organs and/or carcasses and indiscriminate slaughter of pregnant cows represent a significant loss of meat and revenue and a reduction in growth of future herds, which has a negative effect on the livestock industry. This justifies appropriate surveillance and disease control programmes coupled with strict enforcement of legislation governing animal welfare to curb the slaughter of pregnant animals.

Determination of oxytetracycline residues in cattle meat marketed in the Kilosa district, Tanzania.

Oxytetracycline is used to treat various diseases in cattle. However, its use may be associated with unacceptable residue levels in food. Oxytetracycline residues in tissues from indigenous cattle were determined in a cross-sectional study conducted in the Kilosa district, Tanzania, between November 2012 and April 2013. A total of 60 tissue samples, including muscle, liver and kidney, were collected from slaughterhouses and butchers and analysed for oxytetracycline using high-performance liquid chromatography. Oxytetracycline residues were found in 71.1% of the samples, of which 68.3% were above acceptable regulatory levels. The mean concentration of oxytetracycline across tissues was 3401.1 µg/kg ± 879.3 µg/kg; concentrations in muscle, liver and kidney were 2604.1 µg/kg ± 703.7 µg/kg, 3434.4 µg/kg ± 606.4 µg/kg and 3533.1 µg/kg ± 803.6 µg/kg, respectively. High levels of oxytetracycline residue in meat from indigenous cattle may pose a health threat to consumers in Kilosa. The findings possibly reflect a general lack of implementation of recommended withdrawal periods, ignorance about drug use and lack of extension services. Strict regulation of the use of antimicrobial drugs in the livestock industry and associated testing of animal-derived food sources prior to marketing are required.

Assessment of physicochemical characteristics and hygienic practices along the value chain of raw fruit juice vended in Dar es Salaam City, Tanzania.

Fresh fruit juice is an essential component of human diet and there is considerable evidence of health and nutritional benefits. However, nature of the fruits used in juicing and unhygienic processes in the value chain may cause poor quality of juice. This cross- sectional study was conducted to assess physicochemical characteristics and hygienic practices along the value chain of raw fruit juice vended in Dar es Salaam, Tanzania. A total of 90 juice vendors were interviewed. Ninety juice samples were collected and analysed for physicochemical quality. The pH of juices ranged between 2.7 and 6.4, acidity 0.01% and 1.3% and, total soluble solids ranged between -1.5 and 18.04 °Brix. Most juices (67.8%) had -Brix levels below Codex recommended values classified as weak and watery. Juices were made of mango, passion, tamarind, sugar cane and mixture of these fruits sourced from open markets in the city. Water for washing of fruits and dilution of juices was from deep wells (53.3%) and taps (46.7%). About one third (37.8%) of the juice vendors didn't wash the fruits before juicing and 44.4% didn't boil water for juice dilution. Juice extraction was done by kitchen blenders, boiling in water and squeezing by simple machines. Juice pasteurization was not done. The majority of vendors (78.9%) stored juices in plastic buckets and juice was sold in glass cups, reused plastic bottles and disposable cups. Vending sites were restaurants, bus stands and along roadsides. The majority of premises (78.9%) were in unhygienic condition that likely encouraged or introduced contaminants to the juices. It is concluded that, the overall handling, preparation practices and physicochemical quality of raw fruit juices vended in Dare es Salaam City are poor. The government should educate the vendors on food safety and hygiene as well as enforcing regular monitoring of the quality of street fruit juices.

Identification of microcystins in a Lake Victoria cyanobacterial bloom using LC-MS with thiol derivatization.

Microcystins are cyclic heptapeptides from cyanobacteria which are responsible for poisonings of livestock and humans. Cyanobacteria also produce a range of peptides and other compounds that can result in complex chromatograms when samples are analysed by LC-MS. Thiol derivatization of the α,ß-unsaturated amide present in most microcystins was recently shown to simplify analysis of LC-MS chromatograms of a Microcystis culture, making it easier to identify peaks corresponding to microcystins in complex mixtures. This method was applied to analysis of extracts taken from a natural cyanobacteria bloom in Mwanza Gulf, Lake Victoria, Tanzania, in 2010, revealing the presence of numerous putative microcystin analogues in the sample. Results were verified using LC-MS², LC-MS/MS with precursor-ion scanning, and LC-HRMS, leading to identification of 8 major and 17 minor microcystins in the sample, including analogues of microcystin-RY, -RL and -RA. Microcystin-YR (2), -RR (3), and -RY (9) were isolated from bloom material from Lake Victoria, and the structure of 9 was confirmed by NMR spectroscopic analysis and NMR spectral comparison with 2 and 3. Confirmation of the structure of MC-RY (9) facilitated detailed analysis of its MS² spectrum, thereby supporting the structures of related analogues tentatively established on the basis of MS analyses.