Molecular Characterization and Diversity of Bacteria Isolated from Fish and Fish Products Retailed in Kenyan Markets.

Fish products are highly vulnerable to microbial contamination due to their soft tissues, making them perishable and harmful to consumers. The clinical and subclinical infections reported by fish consumers are mainly associated with pathogenic microorganisms in fish products. Therefore, this study aimed at establishing the molecular profiles and diversity of the bacterial isolates from fish and fish products obtained from Kirinyaga County markets in Kenya. A total of 660 samples were randomly sampled in six Kirinyaga County markets and transported to Kenyatta University for bacterial isolation. The fish skin surface was cut using a sterile knife and blended in buffered peptone water. The blended product was serially diluted and plated on nutrient agar. After 24 hours, the bacteria cultures were subcultured to obtain pure bacterial isolates. The pure isolates were grouped and characterized based on their morphology and biochemical characteristics. One representative of each group was selected for bacterial DNA extraction. The 16S rRNA gene was amplified using the 27F and 1492R primers, and the obtained PCR product was subjected to Sanger-based sequencing using the same primers. Morphological characterization yielded 54 morpho groups. Phylogenetic analysis revealed diverse bacterial strains, including Escherichia coli, Salmonella enterica, Citrobacter freundii, Bacillus sp. and Alcaligenes faecalis. Bacillus sp. was the most dominant group, as compared to other isolates in the study. The study, therefore, revealed diverse bacterial strains from the fish products. This high microbial diversity calls for heightened surveillance to prevent possible foodborne disease outbreaks.

Diversity and Phosphate Solubilization Efficiency of Phosphate Solubilizing Bacteria Isolated from Semi-Arid Agroecosystems of Eastern Kenya.

Phosphorus (P) is a major nutrient required for plant growth but it forms complexes with other elements in soil upon application. A cost-effective way of availing P to plants is by use of Phosphate Solubilizing Bacteria (PSB). There is a wide range of PSB suited for diverse agro-ecologies. A large part of Eastern Kenya especially the lower altitude regions are semi-arid with nutrient depleted soils and predominated by low-income smallholders farmers who do not afford costly inorganic fertilizers. To alleviate poor soil nutrition in this agroecosystem, we sought to study the diversity of phosphate solubilizing bacteria and their phosphate solubilization efficiency. The bacteria were selectively isolated in Pikovskaya's agar media. Bacterial colonies were enumerated as Colony Forming Units and morphological characterization determined by analyzing morphological characteristics. Genetic characterization was determined based on sequencing of 16S rRNA. A total of 71 PSB were isolated and they were placed into 23 morphological groups. Correlation analysis showed a negative correlation between phosphate solubilizing bacteria and the levels of phosphorus, iron, calcium, magnesium and soil pH. Analysis of 16S rRNA sequences revealed that the genetic sequences of the isolates matched the strains from the genera Burkholderia, Pseudomonas, Bacillus, Enterobacter, Pantoea, Paraburkholderia, Cronobacter, Ralstonia, Curtobacterium, and Massilia deposited in NCBI Database. Analysis of Molecular Variance showed that variation within populations was higher than that of among populations. Phosphate solubilization index values ranged between 1.143 and 5.883. Findings on biodiversity of phosphate solubilizing bacteria led to identification of 10 candidate isolates for plant growth improvement and subsequently, bio-fertilizer development.

Characterization and diversity of native Azotobacter spp. isolated from semi-arid agroecosystems of Eastern Kenya.

Declining food production in African agroecosystems is attributable to changes in weather patterns, soil infertility and limited farming inputs. The exploitation of plant growth-promoting soil microbes could remedy these problems. Such microbes include Azotobacter; free-living, nitrogen-fixing bacteria, which confer stress tolerance, avail phytohormones and aid in soil bioremediation. Here, we aimed to isolate, characterize and determine the biodiversity of native Azotobacter isolates from soils in semi-arid Eastern Kenya. Isolation was conducted on nitrogen-free Ashby's agar and the morphological, biochemical and molecular attributes evaluated. The isolates were sequenced using DNA amplicons of 27F and 1492R primers of the 16S rRNA gene loci. The Basic Local Alignment Search Tool (BLASTn) analysis of their sequences revealed the presence of three main Azotobacter species viz., Azotobacter vinelandii, Azotobacter salinestris and Azotobacter tropicalis. Kitui County recorded the highest number of recovered Azotobacter isolates (45.4%) and lowest diversity index (0.8761). Tharaka Nithi County showed the lowest occurrence (26.36%) with a diversity index of (1.057). The diversity was influenced by the soil pH, texture and total organic content. This study reports for the first time a wide diversity of Azotobacter species from a semi-arid agroecosystem in Kenya with potential for utilization as low-cost, free-living nitrogen-fixing bioinoculant.

Symbiotic efficiency and genetic characterization of rhizobia and non rhizobial endophytes associated with cowpea grown in semi-arid tropics of Kenya.

Cowpea (Vigna unguiculata (L.) Walp) is an important multipurpose legume crop grown in arid and semi-arid areas of sub-Saharan Africa. The crop associates with a wide diversity of high ecological value rhizobia bacteria, improving biological soil fertility and crop production. Here, we evaluated the symbiotic efficiency (SE) and genetic diversity of native rhizobia isolated from root nodules of cowpea genotypes cultivated in semi-arid areas of lower Eastern Kenya. Rhizobia trapping and SE experiments were done in the greenhouse while genetic diversity was evaluated based on 16S rRNA gene sequencing. Twenty morphologically distinct isolates representing a total of 94 isolates were used for genetic analysis. After 16S rRNA gene sequencing, the isolates closely resembled bacteria belonging to the genus Rhizobium, Paraburkholderia and non-rhizobial endophytes (Enterobacter, Strenotrophomonas and Pseudomonas). This study also reports for the first time the presence of an efficient native cowpea nodulating Beta-Rhizobia (Paraburkholderia phenoliruptrix BR3459a) in Africa. Symbiotic efficiency of the native rhizobia isolates varied (p < 0.0001) significantly. Remarkably, two isolates, M2 and M3 recorded higher SE of 82.49 % and 72.76 % respectively compared to the commercial strain Bradyrhizobium sp. USDA 3456 (67.68 %). Our results form an important step in the development of efficient microbial inoculum and sustainable food production.

Western Kenyan Anopheles gambiae showing intense permethrin resistance harbour distinct microbiota.

BACKGROUND: Insecticide resistance poses a growing challenge to malaria vector control in Kenya and around the world. Following evidence of associations between the mosquito microbiota and insecticide resistance, the microbiota of Anopheles gambiae sensu stricto (s.s.) from Tulukuyi village, Bungoma, Kenya, with differing permethrin resistance profiles were comparatively characterized. METHODS: Using the CDC bottle bioassay, 133 2-3 day-old, virgin, non-blood fed female F1 progeny of field-caught An. gambiae s.s. were exposed to five times (107.5 µg/ml) the discriminating dose of permethrin. Post bioassay, 50 resistant and 50 susceptible mosquitoes were subsequently screened for kdr East and West mutations, and individually processed for microbial analysis using high throughput sequencing targeting the universal bacterial and archaeal 16S rRNA gene. RESULTS: 47 % of the samples tested (n = 133) were resistant, and of the 100 selected for further processing, 99 % were positive for kdr East and 1 % for kdr West. Overall, 84 bacterial taxa were detected across all mosquito samples, with 36 of these shared between resistant and susceptible mosquitoes. A total of 20 bacterial taxa were unique to the resistant mosquitoes and 28 were unique to the susceptible mosquitoes. There were significant differences in bacterial composition between resistant and susceptible individuals (PERMANOVA, pseudo-F = 2.33, P = 0.001), with presence of Sphingobacterium, Lysinibacillus and Streptococcus (all known pyrethroid-degrading taxa), and the radiotolerant Rubrobacter, being significantly associated with resistant mosquitoes. On the other hand, the presence of Myxococcus, was significantly associated with susceptible mosquitoes. CONCLUSIONS: This is the first report of distinct microbiota in An. gambiae s.s. associated with intense pyrethroid resistance. The findings highlight differentially abundant bacterial taxa between resistant and susceptible mosquitoes, and further suggest a microbe-mediated mechanism of insecticide resistance in mosquitoes. These results also indicate fixation of the kdr East mutation in this mosquito population, precluding further analysis of its associations with the mosquito microbiota, but presenting the hypothesis that any microbe-mediated mechanism of insecticide resistance would be likely of a metabolic nature. Overall, this study lays initial groundwork for understanding microbe-mediated mechanisms of insecticide resistance in African mosquito vectors of malaria, and potentially identifying novel microbial markers of insecticide resistance that could supplement existing vector surveillance tools.

Upscaling Arbuscular Mycorrhizal Symbiosis and Related Agroecosystems Services in Smallholder Farming Systems.

Smallholder farming systems form unique ecosystems that can protect beneficial soil biota and form an important source of useful genetic resources. They are characterized by high level of agricultural diversity mainly focused on meeting farmers' needs. Unfortunately, these systems often experience poor crop production mainly associated with poor planning and resource scarcity. Soil fertility is among the primary challenges faced by smallholder farmers, which necessitate the need to come up with affordable and innovative ways of replenishing soils. One such way is the use of microbial symbionts such as arbuscular mycorrhizal fungi (AMF), a beneficial group of soil microbiota that form symbiotic associations with majority of cultivated crops and play a vital role in biological soil fertility, plant nutrition, and protection. AMF can be incorporated in smallholder farming systems to help better exploit chemical fertilizers inputs which are often unaffordable to many smallholder farmers. The present review highlights smallholder farming practices that could be innovatively redesigned to increase AMF symbiosis and related agroecosystem services. Indeed, the future of global food security depends on the success of smallholder farming systems, whose crop productivity depends on the services provided by well-functioning ecosystems, including soil fertility.