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A toxin complex consists of a high-molecular-weight group of toxins that exhibits insecticidal activity against insect pests. These toxins are a promising alternative to Bacillus thuringiensis ( Bt ) toxins that have been extensively utilized in insect pest control. Herein, a codon-optimized insecticidal gene ( tccZ ) (381 bp) identified in Pantoea ananatis strain MHSD5 (a bacterial endophyte previously isolated from Pellaea calomelanos ) was ligated into the pET SUMO expression vector and expressed in Escherichia coli BL21 (DE3). We report the success of cloning the tccZ gene into the pET SUMO vector and ultimately the transformation into E. coli BL21 (DE3) competent cells. However, despite conducting a time course of expression as well as isopropyl β-d-1-thiogalactopyranoside (IPTG) dosage optimization to determine optimal conditions for expression, TccZ protein expression could not be detected on Stain-Free and Coomassie-stained SDS-PAGE gels., Competing Interests: The authors declare that there are no conflicts of interest., (© 2023 The Authors.)

Mahewu is a fermented food product from maize, commonly consumed in Southern Africa. This study investigated the effect of optimizing fermentation (time and temperature) and boiling time of white maize (WM) and yellow maize (YM) mahewu , with the use of the Box-Behnken-response surface methodology (RSM). Fermentation time and temperature as well as boiling time were optimized and pH, total titratable acidity (TTA) and total soluble solids (TSS) determined. Results obtained showed that the processing conditions significantly ( p ≤ 0.05) influenced the physicochemical properties. pH values of the mahewu samples ranged between 3.48-5.28 and 3.50-4.20 for YM mahewu and WM mahewu samples, respectively. Reduction in pH values after fermentation coincided with an increase in TTA as well as changes in the TSS values. Using the numerical multi-response optimisation of three investigated responses the optimal fermentation conditions were observed to be 25 °C for 54 h and a boiling time of 19 min for white maize mahewu and 29 °C for 72 h and a boiling time of 13 min for yellow maize mahewu . Thereafter white and yellow maize mahewu were prepared with the optimized conditions using different inocula (sorghum malt flour, wheat flour, millet malt flour or maize malt flour) and the pH, TTA and TSS of the derived mahewu samples determined. Additionally, amplicon sequencing of the 16S rRNA gene was used to characterise the relative abundance of bacterial genera in optimized mahewu samples, malted grains as well as flour samples. Major bacterial genera observed in the mahewu samples included Paenibacillus , Stenotrophomonas , Weissella , Pseudomonas , Lactococcus , Enterococcus, Lactobacillus, Bacillus , Massilia , Clostridium sensu stricto 1, Streptococcus , Staphylococcus , Sanguibacter , Roseococcus , Leuconostoc , Cutibacterium , Brevibacterium , Blastococcus , Sphingomonas and Pediococcus , with variations noted for YM mahewu and WM mahewu . As a result, the variations in physicochemical properties are due to differences in maize type and modification in processing conditions. This study also discovered the existence of variety of bacterial that can be isolated for controlled fermentation of mahewu .

SCUBA divers are predisposed to otitis externa caused by Pseudomonas aeruginosa, which is becoming increasingly multi-drug resistant (MDR). The present work assessed the antibiotic resistance profiles of P. aeruginosa obtained from SCUBA divers and their environment in Sodwana Bay, South Africa. Bacterial isolates from a total of 137 random water and ear swab samples were identified using biochemical and molecular methods. P. aeruginosa strains were further evaluated for antibiotic susceptibility using the Kirby-Bauer assay. Double disk synergy test (DDST) to confirm metallo-β-lactamase (MBL) production and PCR amplification of specific antibiotic resistance genes was performed. All (100%) 22 P. aeruginosa isolates recovered were resistant to 6 of the β-lactams tested including imipenem but exhibited susceptibility to trimethoprim-sulfamethoxazole. MBL production was observed in 77% of isolates while the most prevalent extended-spectrum β-lactamase (ESBL) genes present included bla AmpC (86.9%) followed by bla TEM (82.6%). Sulfonamide resistance was largely encoded by sul1 (63.6%) and sul2 (77.3%) genes with a high abundance of class 1 integrons (77.3%) of which 18.2% carried both Intl1 and Intl2 . P. aeruginosa found in Sodwana Bay exhibits multi-drug resistance (MDRce) to several pharmaceutically important drugs with the potential to transfer antibiotic resistance to other bacteria if the judicious use of antibiotics for their treatment is not practiced.

This study aimed to investigate the kinetics of phenolic compound modification during the fermentation of maize flour at different times. Maize was spontaneously fermented into sourdough at varying times (24, 48, 72, 96, and 120 h) and, at each point, the pH, titratable acidity (TTA), total soluble solids (TSS), phenolic compounds (flavonoids such as apigenin, kaempferol, luteolin, quercetin, and taxifolin) and phenolic acids (caffeic, gallic, ferulic, p -coumaric, sinapic, and vanillic acids) were investigated. Three kinetic models (zero-, first-, and second-order equations) were used to determine the kinetics of phenolic modification during the fermentation. Results obtained showed that fermentation significantly reduced pH, with a corresponding increase in TTA and TSS. All the investigated flavonoids were significantly reduced after fermentation, while phenolic acids gradually increased during fermentation. Among the kinetic models adopted, first-order (R 2 = 0.45-0.96) and zero-order (R 2 = 0.20-0.82) equations best described the time-dependent modifications of free and bound flavonoids, respectively. On the other hand, first-order (R 2 = 0.46-0.69) and second-order (R 2 = 0.005-0.28) equations were best suited to explain the degradation of bound and free phenolic acids, respectively. This study shows that the modification of phenolic compounds during fermentation is compound-specific and that their rates of change may be largely dependent on their forms of existence in the fermented products.

Lead (Pb) pollution arising from industrial and mining activities has led to widespread environmental toxicity, particularly in South Africa. Humans exposed to Pb are reported to suffer from detrimental health impacts that can lead to fatalities. As such, there is an urgent need to remediate Pb from the environment. In this study, we propose the use of a Pb-specific recombinant fusion metalloprotein, rPbrD surface-cross-linked onto calcium alginate nanoparticles (CANPs) for the biosorption of Pb(II) from aqueous solution. The prepared biosorbents were characterized using scanning electron microscopy, transmission electron microscopy, and dynamic light scattering. Their ability to biosorb soluble Pb(II) was determined by inductively coupled plasma mass spectroscopy and their adsorption mechanism was described according to the Langmuir, Freundlich, Temkin, and Dubinin-Radushkevich adsorption isotherms. The rate of Pb uptake for bare CANPs and rPbrD-CANPs at a concentration of 100 mg/L metal was 3.34 and 8.82 mg/g, respectively, within 30 min. The adsorption data for the bare CANPs best fit the Langmuir isotherm, whereas the adsorption data for rPbrD-CANPs best fitted the Freundlich isotherm. Based on the sorption intensity ( n ) and the separation factor ( R L ), both biosorbents represent a favorable adsorption system. These findings suggest that the proposed nanobiosorbent is a promising candidate for the recovery of Pb ions present in high concentrations such as acid mine drainage or industrial effluent., Competing Interests: The authors declare no competing financial interest., (Copyright © 2019 American Chemical Society.)

PbrD is a lead (II) binding protein encoded by the pbr lead resistance operon found exclusively in Cupriavidus metallidurans CH34. Its ability to sequester Pb(II) shows potential for it to be developed as a biosorbent for Pb in the bioremediation of contaminated wastewaters. In this study the pbrD gene from C. metallidurans CH34 was transformed and overexpressed in Escherichia coli BL21 (DE3) using the pET32 Xa/Lic vector. Optimal expression of recombinant (r)PbrD (∼50 kDa) was achieved post-induction with IPTG within inclusion bodies (IBs). Inclusion bodies were solubilised by denaturation and purified by Ni-NTA affinity chromatography. The purified denatured protein containing the N-terminal Trx•Tag™, His•Tag ® and S ® Tag™ was refolded in vitro via dialysis to a biologically functional form. Circular dichroism spectra of refolded rPbrD-fusion protein indicated a high degree of turns, β-sheets and 3 10 helices content and tryptophan fluorescence showed a structural conformational change in the presence of Pb(II). Refolded rPbrD-fusion protein bound 99.7% of Pb(II) when mixed with lead nitrate in ten-fold increasing concentrations. Adsorption isotherms including Langmuir, Freundlich, Temkin and Dubinin-Radushkevich models were applied to determine the biosorption mechanism. A biologically functional rPbrD-fusion protein has potential application in the development of a biosorbent for remediation of Pb(II) from wastewater., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Enterobacter xiangfangensis Pb204, isolated from acid mine decant from a uranium mine, produces a wide variety of gold nanoparticles (AuNPs), ranging from large triangular plates to small spherical AuNPs. The complete genome sequence of this isolate incorporates an integrative and conjugative element which may be pivotal to AuNP synthesis.

Crimean-Congo hemorrhagic fever (CCHF) is a tick-borne viral zoonosis distributed widely in Africa, Asia, Russia and the Balkans. Occurrence of segment reassortment has been established and may be associated with pathogenicity. The ability to distinguish between an infection with a reassortant and a non-reassortant variant may have prognostic value. In this study the use of Simple-Probe technology and real-time PCR was investigated to detect rapidly CCHF viral nucleic acid and to distinguish between reassorted and non-reassorted variants of southern African CCHF virus (CCHFV) isolates. A Simple-Probe was designed based on multiple alignments of 19 CCHFV partial M segment sequences that included reassorted and non-reassorted isolates from two phylogenies. Real-time PCR followed by probe-specific melting-curve analysis allowed differentiation of three selected isolates representative of two phylogenetically distinct groups, SPU 431/85 (group IV), 383/87 (group IV) and 103/87 (group III) based on melting temperatures. Within group IV there are two distinct lineages which were represented in the optimization. A further 16 clinical isolates could be genotyped into groups III and IV using the Simple-Probe real-time PCR assay developed in this study. This assay provides a reliable and sensitive tool for the differentiation between reassorted and non-reassorted variants of CCHFV which finds application for diagnosis and epidemiologic and surveillance studies., (Copyright (c) 2010 Elsevier B.V. All rights reserved.)

Beak and feather disease virus (BFDV), the causative agent of psittacine beak and feather disease (PBFD) infects psittaciformes worldwide. We provide an annotated sequence record of three full-length unique genomes of BFDV isolates from budgerigars (Melopsittacus undulatus) from a breeding farm in South Africa. The isolates share >99% nucleotide sequence identity with each other and approximately 96% nucleotide sequence identity to two recent isolates (Melopsittacus undulatus) from Thailand but only between 91.6 and 86.6% identity with all other full-length BFDV sequences. Maximum-likelihood analysis and recombination analysis suggest that the South African budgerigar BFDV isolates are unique to budgerigars, are non-recombinant in origin, and represent a new genotype of BFDV.

Psittacine beak and feather disease (PBFD) is a common viral disease of wild and captive psittacine birds characterized by symmetric feather loss and beak deformities. The causative agent, beak and feather disease virus (BFDV), is a small, circular single-stranded DNA virus that belongs to the genus Circovirus. BFDV can be detected by PCR or the use of haemagglutination (HA) and haemagglutination inhibition (HI) assays that detect antigen and antibodies respectively. Erythrocytes from a limited number of psittacine species of Australian origin can be used in these tests. In South Africa, the high cost of these birds makes them difficult to obtain for experimental purposes. Investigation into the use of erythrocytes from African Grey parrots and Brown-headed parrots yielded positive results showing the haemagglutinating activity of their erythrocytes with purified BFDV obtained from confirmed clinical cases of the disease. The HA activity was further confirmed by the demonstration of HI using BFDV antiserum from three different African Grey parrots previously exposed to the virus and not showing clinical signs of the disease.