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Lignocellulose constitutes a major component of discarded wastes from various industries viz. agriculture, forestry and municipal waste treatment. The potential use of lignocellulose from such types of biomass can be maximized by enzymatic degradation using glycoside hydrolases (GHs) and oxidative enzymes to produce renewable fuels. Nonetheless, besides the slow rate of degradation and low yields, lignocellulose is also physicochemically recalcitrant and costly to process, further limiting its mass utilization. Therefore, bioprospecting for microorganisms producing efficient lytic polysaccharide monooxygenases (LPMOs) to overcome these drawbacks may prove beneficial. The use of GHs and LPMOs can potentially help to circumvent some limitations in the conversion of lignocellulosic biomass into fermentable sugars. LPMOs are classified as family GH61 or family 33 carbohydrate-binding module (CBM33), whose unusual surface-exposed active site is bound to a copper (II) ion. To date, there are more than 20 known genes encoding cellulose-active LPMOs in bacteria and fungi, with diverse biological activities. Only by thorough comprehension of the diversity, enzymology and role of primary GHs, i.e. celullases and their oxidative machinery can the degradation of lignocellulosic biomass be improved. This review provides insight into the diversity, structure and mechanisms, structural and functional aspects of the oxidative breakdown of cellulose by LPMOs of the cellulose-active GH family.

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BACKGROUND:The growing demand for cellulases and xylanases in lignocellulosic degradation and reutilization has spurred the need for their improved production at reduced cost. Abundantly available oil palm leaves (OPL) promise an alternative and sustainable substrate for producing such enzymes in the degradation and biotransformation of unwanted lignocellulosic biomass. Both enzymes are key players in the transformation of biomass into other value-added commodities.RESULTS: This study statistically optimized the solid-state fermentation (SSF) of raw OPL by novel fungal strains Trichoderma asperellum UC1 and Rhizopus oryzae UC2, to produce cellulase and xylanase, where raw OPL was pretreated using ultrasonication before the dual enzyme-assisted saccharification process. This experiment aimed to identify the best parameters for the highest activity of CMCase (EC 3.2.1.4), FPase (EC 3.2.1.91) and xylanase (EC 3.2.1.8) of T. asperellum UC1 and of ⊎-glucosidase (EC 3.2.1.21) of R. oryzae UC2 for improved saccharification. Activities of CMCase (126.87 U g −1 ), FPase (85.53 U g −1 ) and xylanase (215.42 U g −1 ) achieved the maximum under optimized SSF conditions (30 °C, 2.0 × 10 7 spores g −1 , 75% moisture content, pH 6). The best ⊎-glucosidase activity (131.76 U g −1 ) was obtained at 32 °C, 2.0 × 10 7 spores g −1 , 50% moisture content, pH 12. OPL saccharification yielded 1240 ± 32 mg g −1 total reducing sugar. Individual enzyme cocktails improved juice clarification (84-88%) and dough rising (1.7-to 2.0-fold).CONCLUSIONS: The optimized SSF of raw OPL successfully afforded the production of effective cellulases and xylanases for saccharification-related reactions.

Aim: This study was aimed to determine the virulent genes and antibiotic resistance patterns among circulating diarrheagenic Escherichia coli (DEC) pathotypes in a tertiary care health center in east of Nigeria. Materials and Methods: Diarrheal stool samples were obtained from 80 children under 5 years and E. coli was isolated and identified using standard biochemical and molecular methods. Multiplex polymerase chain reaction (PCR) was used to detect eight virulent genes of DEC. Disk diffusion method was used to determine the antibiotic susceptibility of DEC. Results: DEC infection was observed in 54 (68%) children among which ial gene for enteroinvasive E. coli (EIEC) (40% [n=22]) was commonly detected followed by eltA/eltB for enterotoxigenic E. coli (ETEC) (30% [n=16]), pCVD for enteroaggregative E. coli (EAEC) (20% [n=11]), and eaeA/bfpA for typical enteropathogenic E. coli (EPEC) (10% [n=5]). The DEC isolates phenotypically exhibited resistance for ampicillin (AMP) (44 [81%]), followed by ciprofloxacin (CIP)/ levofloxacin (LEV) (28 [52%]), cefoxitin (FOX) (11 [20%]), and amoxicillin-clavulanic acid (AMC) (6 [11%]). About 60% isolates of stable toxins-ETEC were resistant to AMC, CIP, and LEV while all the labile toxin-ETEC exhibited resistance to AMP. About 60% (n=6) resistance were seen in EAEC against ampicillin, AMC, FOX, CIP, and LEV. In EIEC, all the isolates (n=22) were resistant to AMP while 50% (n=11) were resistant to both CIP and LEV. All EPEC (n=5) were resistant to AMP, FOX, CIP, and LEV. Conclusion: High frequency of virulent ial and eltA/eltB genes for EIEC and ETEC, respectively, suggests that they are the primary etiological agents of diarrhea in children among DEC pathotypes. Resistance of DEC to more than two classes of antibiotics indicate possible emergence of multidrug resistance.

Background. Diarrheagenic E. coli (DEC) is an etiological agent of childhood diarrhea. Resistance against commonly used drugs in the empirical treatment of enteric infections has increased among DEC. Relationship between antibiotic resistance and biofilm formation in microorganisms have been widely reported. This study was aimed to determine the antibiotic resistance and biofilm production pattern among DEC pathotypes isolated from stools of children aged 0–5 years with acute diarrheal disease in Abakaliki, Nigeria. Materials and methods. Diarrheal stool samples were obtained from 60 children and E. coli were isolated and identified using standard guidelines provided for laboratory diagnosis of enteric pathogens. Molecular identification was done by amplification of E. coli universal stress protein A (uspA) using polymerase chain reaction (PCR) method. Detection of virulent genes of DEC pathotypes was performed in a group of multiplex PCR using their specific primers. Kirby–Bauer disk diffusion method was used to determine the antibiotic susceptibility patterns of the isolates while biofilms production was detected by thiazolyl blue tetrazolium bromide dye in a 96-well plate. Results. DEC was isolated in 40 stools among which EIEC [40% (n = 16)] was commonly detected followed by ETEC [30% (n = 12)], EAEC [20% (n = 8)] and typical EPEC [10% (n = 4)]. Half of EAEC showed the highest multidrug resistance against ampicillin, cefoxitin, ciprofloxacin, levofloxacin, and tetracycline with the strongest biofilm production followed by all the EPEC which were resistant to ampicillin, ciprofloxacin, levofloxacin, and tetracycline with moderate biofilm production. All the LT-ETEC exhibited the least resistance to ampicillin and tetracycline with the weakest biofilm production. Conclusion. High frequency of the EIEC pathotype suggests its role as the primary etiological agent of diarrhea in children. Correlation between high drug resistance and biofilm production among the pathotype may indicate that biofilms may provide compatible uptake of resistance genes.

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