Paenibacillus Spp Research Articles

Paenibacillus spp infection among infants with postinfectious hydrocephalus in Uganda: an observational case-control study

Paenibacillus thiaminolyticus is a cause of postinfectious hydrocephalus among Ugandan infants. To determine whether Paenibacillus spp is a pathogen in neonatal sepsis, meningitis, and postinfectious hydrocephalus, we aimed to complete three separate studies of Ugandan infants. The first study was on peripartum prevalence of Paenibacillus in mother-newborn pairs. The second study assessed Paenibacillus in blood and cerebrospinal fluid (CSF) from neonates with sepsis. The third study assessed Paenibacillus in CSF from infants with hydrocephalus. In this observational study, we recruited mother-newborn pairs with and without maternal fever (mother-newborn cohort), neonates (aged ≤28 days) with sepsis (sepsis cohort), and infants (aged ≤90 days) with hydrocephalus with and without a history of neonatal sepsis and meningitis (hydrocephalus cohort) from three hospitals in Uganda between Jan 13, 2016 and Oct 2, 2019. We collected maternal blood, vaginal swabs, and placental samples and the cord from the mother-newborn pairs, and blood and CSF from neonates and infants. Bacterial content of infant CSF was characterised by 16S rDNA sequencing. We analysed all samples using quantitative PCR (qPCR) targeting either the Paenibacillus genus or Paenibacillus thiaminolyticus spp. We collected cranial ultrasound and computed tomography images in the subset of participants represented in more than one cohort. No Paenibacillus spp were detected in vaginal, maternal blood, placental, or cord blood specimens from the mother-newborn cohort by qPCR. Paenibacillus spp was detected in 6% (37 of 631 neonates) in the sepsis cohort and, of these, 14% (5 of 37 neonates) developed postinfectious hydrocephalus. Paenibacillus was the most enriched bacterial genera in postinfectious hydrocephalus CSF (91 [44%] of 209 patients) from the hydrocephalus cohort, with 16S showing 94% accuracy when validated by qPCR. Imaging showed progression from Paenibacillus spp-related meningitis to postinfectious hydrocephalus over 1-3 months. Patients with postinfectious hydrocephalus with Paenibacillus spp infections were geographically clustered. Paenibacillus spp causes neonatal sepsis and meningitis in Uganda and is the dominant cause of subsequent postinfectious hydrocephalus. There was no evidence of transplacental transmission, and geographical evidence was consistent with an environmental source of neonatal infection. Further work is needed to identify routes of infection and optimise treatment of neonatal Paenibacillus spp infection to lessen the burden of morbidity and mortality. National Institutes of Health and Boston Children's Hospital Office of Faculty Development.

Large hospital-wide outbreak of Paenibacillus spp pseudobacteremia associated with contaminated nonsterile gloves.

We report a large, hospital-wide outbreak of pseudobacteremia by Paenibacillus spp. In total, 139 patients presented at least 1 positive blood culture during a 13-month period. Microbiological experiments indicated that contaminated nonsterile gloves were associated with the pseudobacteremia episodes. The outbreak was resolved by discontinuing the use of the involved brand.

Characterization of Paenibacillus spp. CBRM17 as antagonist of phytopathogenic fungi and growth promoter of Capsicum chinense Jacq.

Studies of the plant-microorganism relationship have made it posible to explore the potential of rhizospheric bacteria to improve the health and quality of plants. In the present study, Paenibacillus sp. CBRM17 was characterized in vitro for its ability to inhibit the growth of phytopathogenic fungi that cause diseases in horticultural crops in the tropics, finding a reduction in the mycelial growth of Alternaria, Fusarium and Helmintosphorium strains in the range of 50 to 70 %, additionally its biochemical properties related to the promotion of plant growth were characterized, registering the production of 0.36 µg/mL of indole acetic acid (IAA), the solubilization of tricalcium phosphate with solubilization index (SI) of 2.41 mm and solubilization efficiency (SE) of 140.6 %, producing in the supernatant 63.5 mg/L of soluble phosphorus, in addition to being positive for ACC deaminase activity. Inoculations trials of habanero chili (Capsicum chinense Jacq.) seeds with Paenibacillus sp. CBRM17 showed its potential to be used as an inoculant in the growth promotion of this crop, since it increased all growth variables; increasing the total fresh and dry biomass by 93.3 and 96.4 %, respectively.

Microbiological Assessment of Used Face Masks in Port Harcourt Secondary Schools in COVID-19 Era

Aim: This study was aimed at identifying bacterial and fungal contaminations in used face masks from different secondary schools in Port Harcourt during COVID-19 Era
 Study Design: The study employs statistical analysis of the data and interpretation.
 Place and Duration of Study: Five Secondary Schools–Three public schools: Federal Government College Rumuokoro; Rumueme and Rumuokuta Girls’ Secondary Schools; Two Private Schools: Solid Steps and Istan Secondary Schools; all located in the city of Port-Harcourt, Nigeria. Sample collection lasted for a week and the analysis lasted for six months.
 Methodology: The research study was facilitated through Laboratory analysis and the use of questionnaire to get the age and sex from the school children. A total of 25 used face masks samples were collected from school children between ages of 12-18years and they were examined microbiologically. Sterile swab sticks soaked in sterile nutrient broth were employed to swab the inner surface area of the used face mask of circular diameter 10 cm. The swabbed samples were dipped and shaken in 9ml of sterile saline water for 1-3 minutes to dislodge the organisms; the mixture was then diluted through a ten-fold serial dilution, after which an aliquot of 0.1ml were inoculated unto Nutrient Agar (dilution used 10-6, incubated at 370C for 24h), Mac Conkey Agar (dilution used 10-3, incubated at 44±0.20C for 24-48h) and Sabouraud Dextrose Agar (dilution used 10-3; incubated at 370C for 5-7 days). Frequency evaluation and identification of isolates were carried out using standard microbiological techniques.
 Results: The entire face masks sampled were found contaminated with microorganisms. The Microbial load (Log10 CFU/cm2;) and Percentage (%) occurrence of bacterial isolates from used facemask were; Bacillus spp (6.10±2.13)(30.81) > Staphylococcus auerus (3.89±3.01)(19.57%) > Proteus spp (2.25±2.45)(11.35) > Paenibacillus spp (1.55±2.52)(7.82) > Escherichia coli (0.36±0.81)(1.82) while fungal isolates were Aspergillus spp (2.20±0.55)(11.09) > Mucor spp (2.19±0.96)(11.04) > Penicillum spp (1.29±0.61)(6.51). The contaminated used face masks with microorganism were highest in school children of ages 16-18years (72%) and the lowest occurred in children of 12-14years of age (12%).
 Conclusion: The presence of potential pathogen such as Staphylococcus auerus, Bacillus spp etc. are of public health significance. It is therefore recommended that crowd should be controlled in such environments with high bacterial and fungal load such as schools and COVID-19 protocols duly observed.

Bio-Organic Fertilizer: A Green Technology to Reduce Synthetic N and P Fertilizer for Rice Production.

Decomposed organic materials, in combination with plant growth-promoting bacteria (PGPB), are environmentally friendly and reduce synthetic fertilizer use in rice production. A bio-organic fertilizer (BoF) was prepared using kitchen waste (79%), chita-dhan (unfilled rice grain) biochar (15%), rock phosphate (5%), and a consortium of 10 PGPB (1%) to supplement 30% nitrogen and to replace triple superphosphate (TSP) fertilizer in rice production with an improvement of soil health. PGPB were local isolates and identified using 16S ribosomal RNA partial gene sequences as Bacillus mycoides, Proteus sp., Bacillus cereus, Bacillus subtilis, Bacillus pumilus, Paenibacillus polymyxa, and Paenibacillus spp. Isolates could fix N2 by 0.7–1.4 g kg–1, solubilize 0.1–1.2 g kg–1 phosphate, and produce 0.1–40 g kg–1 indoleacetic acid. The performance of BoF was evaluated by 16 field experiments and 18 farmers’ field demonstration trials during the year 2017–2020 in different parts of Bangladesh. Performances of BoF were evaluated based on control (T1), full synthetic fertilizer dose of N, P, and K (T2), BoF (2 t ha–1) + 70% N as urea + 100% K as muriate of potash (T3), 70% N as urea + 100% P as TSP + 100% K as muriate of potash (T4), and 2 t ha–1 BoF (T5) treatments. At the research station, average grain yield improved by 10–13% in T3 compared with T2 treatment. Depending on seasons, higher agronomic N use efficiency (19–30%), physiological N use efficiency (8–18%), partial factor productivity (PFP)N (114–150%), recovery efficiency (RE)N (3–31%), N harvest index (HIN) (14–24%), agronomic P use efficiency (22–25%), partial factor productivity of P (9–12%), AREP (15–23%), and HIP (3–6%) were obtained in T3 compared with T2 treatment. Research results were reflected in farmers’ field, and significant (P < 0.05) higher plant height, tiller, panicle, grain yield, partial factor productivity of N and P were obtained in the same treatment. Application of BoF improved soil organic carbon by 6–13%, along with an increased number of PGPB as compared with full synthetic fertilizer dose. In conclusion, tested BoF can be considered as a green technology to reduce 30% synthetic N and 100% TSP requirements in rice production with improved soil health.

Spore Heat Resistance and Growth Ability at Refrigeration Temperatures of Bacillus spp. and Paenibacillus spp.

In this study, spore heat resistance and growth ability at refrigeration temperatures of Bacillus spp. and Paenibacillus spp. were determined. The spore D90°C of 67.6% (23 of 34 strains) of Bacillus and 73.9% (17 of 23 strains) of Paenibacillus was less than 15 min. The growth abilities of both genera were equivalent at 10°C. However, 71.1% (32 of 45 strains) of Paenibacillus and only 6.3% (3 of 48 strains) of Bacillus cereus group could grow at 4°C. Eight B. cereus strains formed spores with higher heat resistance compared to the other Bacillus strains assessed; however, they did not grow at tempreratures below 10°C. Conversely, four Paenibacillus strains formed spores with heat resistance equivalent to that of the eight B. cereus strains and grew at 6°C or lower. In particular, Paenibacillus sp. JCM13343 formed the highest heat-resistant spores (D90°C = 136.1 min) and grew well at 4°C. These results indicate that Paenibacillus can grow in processed foods during refrigerated storage and has the potential to cause spoilage as well as Bacillus. Therefore, Paenibacillus should be considered as one of the targets for microbiological control in refrigerated processed foods.

Management of microbial contaminants in the In Vitro Gene Bank: a case study of taro [Colocasia esculenta (L.) Schott

Ex situ conservation of vegetatively propagated crops by tissue culture is sometimes hampered by covert endophytic bacteria which become visible after repeated subculture over prolonged periods. In the present study, we identified bacterial contaminants and devised a strategy for their elimination from 20 in vitro conserved accessions of taro [Colocasia esculenta (L.) Schott] held in the In Vitro Gene Bank (IVGB) at ICAR-National Bureau of Plant Genetic Resources (NBPGR). Visually, the cultures were exhibiting white to dark brown exudation in tissue culture growth medium and undergoing slow degeneration as recorded by wilting and necrosis of leaves and shoots. On culturing the macerated pieces of shoots, corms, and leaf petioles from the contaminated taro cultures on bacterial indexing medium, six distinct bacterial colonies were observed. The identity of these bacteria was established through 16S rDNA sequencing as gram-negative strains (2) of Ralstonia spp. and gram-positive strains (4) of Paenibacillus spp. Thereafter, two strategies were adopted for elimination of contaminants from the cultures: (i) use of antibiotic supplemented media and (ii) hardening of micro-corms in field and re-establishment under in vitro conditions. Antibiotic supplemented media was not effective in eliminating contamination as reappearance of bacterial colonies was observed after subculturing in antibiotic-free growth medium. In contrast, re-establishment in vitro from hardened infected plants resulted in an average of 6.03% bacteria-free cultures in 14 accessions. The findings indicate that endophytic bacteria associated with the host (taro) plants under tissue culture conditions may efficiently be mitigated through periodic transfer of the tissue culture derived corms to ex vitro conditions and re-introduction of the bacteria-free shoot tip explants in vitro.

In vitro assessment of some probiotic properties and inhibitory activity against Yersinia ruckeri of bacteria isolated from rainbow trout Oncorhynchus mykiss (Walbaum)

In the present study, we isolated bacteria from intestines of healthy rainbow trout, screened for their probiotic properties, and inhibitory activity against Y. ruckeri. A total of 21 out of 541 isolates from the intestine of adult rainbow trout (∼ 200–350 g), showed inhibition zones around at least one Y. ruckeri strains tested. Six of them were selected due to their ability to inhibit all the pathogenic strains on solid medium. They were identified as B. amyloliquefaciens 131and Paenibacillus spp (code 134, 1cc, 1d, 1k, and 2cc). Excluding the isolate 1k, selected strains were able to inhibit Y. ruckeri strains at 22 and 15 ± 0.5 °C when they were simultaneously cocultured, however, they did not show inhibition at 12 ± 0.5 °C. A pre incubation process of the selected strains for 24 h prior to co-culture with strains of Y. ruckeri enabled them to inhibit target pathogens at the lowest temperature tested. Furthermore, probiotic candidates survived to high concentrations of rainbow trout bile salts and acidic conditions. Selected bacteria were also able to produce amylolitic, proteolytic and lipolytic enzymes and showed variable hydrophobic values. Biosafety assays, performed by oral and intraperitoneal infection using B. amyloliquefaciens 131 and Paenibacillus spp (code 134, 1cc, 1d, 1k, and 2cc), showed no pathogenic effect in rainbow trout during 20-day experiment. Therefore, selected strains showed potential characteristics that are required to be evaluated in vivo for their possible subsequent use as probiotics in rainbow trout aquaculture.

Plant growth promotion and alleviation of salts in rice by Paenibacillus spp

Soil salinity plays an evil role by making a crop highly fragile and thereby deteriorating its productivity. Rice is a salt sensitive crop and soil salinity is the single most widespread soil toxicity problem facing rice production. Major problem soils under which rice is grown are Sodic soils, Saline soils (includes inland and coastal saline soils) and Saline-sodic soils and these soils adversely affect growth and productivity of rice. In this study, Paenibacillus castaneae VPB1 and Paenibacillus stellifer KVPB5 were evaluated for alleviating salt stress. The isolates could tolerate salinity stress up to 15% NaCl. The isolates possessed traits like nitrogen fixation, phosphorous solubilisation, siderophore and IAA production. The two isolates were taken for gnotobiotic assay under salt stressed conditions. A considerable growth of rice seedling (CO-51) was noted under gnotobiotic assay. The pot culture studies were performed under different salt stress (0 mM NaCl, 50 mM NaCl, 100 mM NaCl). Among various treatments, the treatment T5 (Paenibacillus castaneae VPB1 + 50 mM NaCl) recorded maximum shoot length of 19.7 and 56.0 cm and root length of 18.6 and 37.5 cm at 30 and 60 DAS respectively. Paenibacillus castaneae VPB1 plus 50 mM NaCl significantly increased the number of grains (201 plant-1) treatment without bacterial inoculation with salt stress. Hence, it is concluded that Paenibacillus spp. could be recommended as potential bio-inoculant to rice crop under salt stressed conditions after field evaluation.

Development of a multiplex real-time PCR assay for the identification and quantification of group-specific Bacillus spp. and the genus Paenibacillus

Spoilage microorganisms can occur at many points throughout food production systems. Bacillus spp. and Paenibacillus spp. are important aerobic spoilage bacteria in various sectors of the food industry. In this study, we developed a rapid detection and quantification technique for Bacillus group-specific and the genus Paenibacillus by using multiplex quantitative PCR (mqPCR). The 1st was the Bacillus cereus group containing B. cereus and B. weihenstephanensis; the 2nd was the B. subtilis group containing B. subtilis, B. licheniformis, B. safensis, and B. pumilus; the 3rd was the B. simplex group containing B. megaterium and B. simplex; and the 4th was the genus Paenibacillus. Depending on the assays, the detection limit was 10 copy numbers. In addition, mqPCR assays were validated by spiking potato salad and milk samples with four strains; B. weihenstephanensis, B. licheniformis, B. megaterium, and P. lautus. The detection dynamic range for potato salad was 105 CFU/mL–101 CFU/mL with B. weihenstephanensis and B. licheniformis, and 105 CFU/mL–102 CFU/mL with B. megaterium and P. lautus, while, for milk, all strains were 105 CFU/mL–102 CFU/mL. We also stored these food matrices spiked with four bacterial suspensions (approximately 103 CFU/mL) at various temperatures. Results showed that B. weihenstephanensis and B. licheniformis were able to grow in potato salad, whereas, the populations of B. weihenstephanensis, B. licheniformis, and P. lautus increased in milk. Consequently, the mqPCR assays developed here in facilitated the differentiation, quantification, and confirmation of the presence of the psychrophilic and psychrotolerant Bacillus group and Paenibacillus spp.

2311. Bacteremia Is Not Commonly Detected in Ebola Virus Disease

BackgroundRates of bacteremia in Ebola virus disease (EVD) are not currently known. Given the potential for secondary bacterial infection during acute EVD, current treatment guidelines recommend empiric use of broad-spectrum antibiotics. We sought to determine rates of bacteremia among patients evaluated for EVD at the ELWA-3 Ebola Treatment Unit in Monrovia, Liberia during the 2013–16 West Africa epidemic.MethodsDeidentified blood samples and matched clinical data from 235 Ebola virus (EBOV)-positive patients and 102 EBOV-negative patients were evaluated under a University of Liberia Pacific Institute for Research and Evaluation IRB-approved protocol. 0.2 mL aliquots of frozen whole blood samples collected at triage, prior to the administration of antibiotics, were inoculated into BD BACTEC Peds Plus bottles and incubated under aerobic conditions in a BD BACTEC FX40 for 5 days in the National Institute of Allergy and Infectious Disease Biosafety Level 4 laboratory in Hamilton, MT. Positive samples were sub-cultured on nonselective sheep blood agar and chocolate agar and pure colonies were selected for identification by 16S sequencing and by matrix assisted laser desorption ionization time-of-flight mass spectrometry.ResultsNo difference in rates of bacteremia was detected among EBOV-positive vs. EBOV-negative patients – 3.8% and 3.9%, respectively. Predominant isolates included Staphylococcus epidermidis and other coagulase-negative staphylococci, thought consistent with contaminants. Pathogenic species included Staphylococcus aureus and possibly Paenibacillus spp.ConclusionThese data suggest that bacteremia does not commonly complicate EVD. However, as both prolonged sample storage and low culture volume may negatively affect sensitivity, additional molecular analyses are needed to support this conclusion.The Intramural Research Program of the National Institutes of Health supported this work.Disclosures All authors: No reported disclosures.

Efficient Reduction of Iron Oxides by Paenibacillus spp. Strains Isolated from Tropical Soils

The genus Paenibacillus was hardly described as a Fe(III)-reducing agent, only limited to reduce soluble forms or Fe inserted in poorly crystallized structures. In this study, three Paenibacillus strains capable of reducing manganese oxides in addition to iron oxides were isolated from Cameroonian and Brazilian soils. These strains reduced iron minerals from poorly crystallized 2-line ferrihydrite to well-crystallized Al-substituted and pure goethite with a significant production of soluble ferrous iron. These Paenibacillus strains, inhabitants from ferralitic temporarily waterlogged soils, could play an important role in the bioweathering of minerals and metal mobility in soils.

Natural Attenuation Potential of Polychlorinated Biphenyl-Polluted Marine Sediments

The marine environment in Kuwait is polluted with various hazardous chemicals of industrial origin. These include petroleum hydrocarbons, halogenated compounds and heavy metals. Bioremediation with dedicated microorganisms can be effectively applied for reclamation of the polluted marine sediments. However, information on the autochthonous microbes and their ecophysiology is largely lacking. We analyzed sediments from Shuwaikh harbor to detect polychlorinated biphenyls (PCBs) and total petroleum hydrocarbons (TPHs). Then we adopted both culture-dependent and culture-independent (PCR-DGGE) approaches to identify bacterial inhabitants of the polluted marine sediments from Shuwaikh harbor. The chemical analysis revealed spatial variation among the sampling stations in terms of total amount of PCBs, TPHs and the PCB congener fingerprints. Moreover, in all analyzed sediments, the medium-chlorine PCB congeners were more abundant than the low-chlorine and high-chlorine counterparts. PCR-DGGE showed the presence of members of the Proteobacteria, Spirochaetes, Firmicutes and Bacteroidetes in the analyzed sediments. However, Chloroflexi-related bacteria dominated the detected bacterial community. We also enriched a biphenyl-utilizing mixed culture using the W2 station sediment as an inoculum in chemically defined medium using biphenyl as a sole carbon and energy source. The enriched mixed culture consisted mainly of the Firmicute Paenibacillus spp. Sequences of genes encoding putative aromatic ring-hydroxylating dioxygenases were detected in sediments from most sampling stations and the enriched mixed culture. The results suggest the potential of bioremediation as a means for natural attenuation of Shuwaikh harbor sediments polluted with PCBs and TPHs.

Effect of oxygen supply on milk-clotting activity expressed by Paenibacillus spp. strain BD3526

Paenibacillus spp. BD3526, a novel bacteria species isolated from raw yak (Bos grunniens) milk collected in Tibet, is able to produce metalloproteinase with milk-clotting activity (MCA) in wheat bran broth. To determine the critical factor favoring enzyme productivity, response surface methodology and the Box-Behnken design were adopted to preliminarily optimize BD3526 cultivation conditions for maximum MCA in shaking flasks. The highest degree of MCA (7072 ± 519 SU/mL) was obtained at a liquid volume of 30 mL and 300 r/min, which implies that oxygen supply is crucial for enzyme synthesis. MCA levels were enhanced as agitation speeds and aeration rates were increased in a 7-L scale fermentor. MCA in the supernatant was further boosted by adding hydrogen peroxide (H2O2) to the broth during cultivation. Maximum MCA (14778 ± 1100 SU/mL) and MCA/proteolytic activity (PA) (33000 ± 918) were achieved when cultivation was carried out at an agitation speed of 400 r/min, aeration rate of 5.5 L/min and H2O2 flow of 500 mmol/L at a feeding rate of 2.5 mL/h after 24 h of cultivation.

Structural insight into a novel neutral metalloproteinase from Paenibacillus spp. BD3526: Implications for mechanisms of rapid inactivation and calcium-dependent stability

The proteinase with milk-clotting activity (MCA) from Paenibacillus spp. BD3526 is characterized as a neutral metalloproteinase of 35kDa. However, the rapid reduction of its MCA during separation and purification leads to low enzyme recovery. The effects of metal ions, inactivation kinetics, and concentration of calcium on the enzymatic activities and thermal stability of the BD3526 metalloproteinase were investigated. In the absence of calcium, the residual activities of the BD3526 metalloproteinase sharply declined during the first three hours, and continued to slowly decrease thereafter. The activities were well fitted by a double-exponential decay model. The inactivation rates were significantly inhibited by calcium and the residual enzyme activities were maintained at more than 80% for 30d at room temperature with 50–100mM calcium. An intermolecular autoproteolytic mechanism was responsible for BD3526 metalloproteinase inactivation. The target protein band with MCA remained largely undegraded in the enzyme solution that was supplemented with 100mM of calcium, but gradually diminished over time in the absence of calcium. N-terminal amino acid sequencing showed that cleavage at the His252-Ala253 peptide bond facilitated the conversion of the zymogen into the active enzyme. Sequence alignment revealed the presence of two highly conserved motifs, HEXXH and GXXNEXXSD, indicating that the enzyme belonged to the metalloproteinase family M4, also known as thermolysin-like proteinases (TLPs). Further structural analysis showed that the observed calcium-dependent stability of the BD3526 TLP may be attributable to a partly degenerated calcium-binding site, Ca1-2, and a mutant calcium-binding site, Ca3.

Medium optimization for nitrogen fixer Paenibacillus sp. 1-49.

Paenibacillus sp. 1-49 is a nitrogen-fixing bacterium that has the potential as a fertilizer. However, it grows poorly (OD600≤1) in both mineral medium and rich medium. To achieve high-yield biomass, we optimized fermentation medium for Paenibacillus sp. 1-49. Plackett-Burman Design and Central Composite Design in response surface methodology were used to optimize medium composition. The optimal fermentation medium contained:(per liter) 36.22 g Sucrose, 5.31 g Tryptone, 10.92 g Yeast Extract, 0.51 g MgSO4, 3.5 g NaCl, 0.02 g Na2MoO4 and 0.02 g FeSO4. The maximum OD600 of 10.280±0.009 was obtained in shake flask fermentation, reached 94.6% of the predicted value. We succeeded in using a response surface methodology to optimize the fermentation medium for Paenibacillus sp. 1-49. Our results will be useful for largescale fermentation of this strain and other Paenibacillus spp.

Concurrence of spores of Clostridium tyrobutyricum, Clostridium beijerinckii and Paenibacillus polymyxa in silage, dairy cow faeces and raw milk

Spores of lactate-fermenting clostridia, known as butyric acid bacteria (BAB), can cause severe quality defects in semi-hard cheeses, called late-blowing. The routine detection method of BAB spores involves most probable number quantification of spores of anaerobic, gas-forming bacteria in a medium containing lactic acid. In this study, BAB spores were detected in 296 samples of soil, maize and grass silage, dairy cow faeces and farm tank milk collected from dairy farms in The Netherlands and the most abundant populations identified. Three major populations were detected, namely Clostridium tyrobutyricum, Clostridium beijerinckii and Paenibacillus spp. Paenibacillus polymyxa was the most abundant species of the latter group. The results indicate that the three spore populations share the same sources and the same contamination route of milk. In contrast to the Clostridium species, P. polymyxa isolates were unable to ferment lactate into butyrate. P. polymyxa spores are presumably unable to cause cheese defects.

Purification and characterization of a novel milk-clotting metalloproteinase from Paenibacillus spp. BD3526

In this study, a milk-clotting enzyme (MCE) isolated from Paenibacillus spp. BD3526 was purified and characterized. The MCE was purified 8.9-fold with a 10.11% recovery using ammonium sulfate precipitation and anion-exchange chromatography and the specific milk-clotting activity (MCA) reached 6791.73SU/mg. The enzyme was characterized as a 35kDa metalloproteinase, and the zymogen of which was encoded by a 1671bp gene named zinc metalloproteinase precursor (zmp) with a predicted molecular weight of 59.6kDa. The optimal temperature for MCA and proteolytic activity (PA) was 65°C and 60°C, respectively. The enzyme was stable over a pH range of 5.0–9.0 and at temperatures below 50°C. The MCA was completely inactivated when the enzyme was heated at 60°C for 30min, and the PA was totally inactivated for 20 and 10min when the enzyme was heated at 55°C and 60°C, respectively. The BD3526 enzyme was preferentially active towards κ-casein (κ-CN) and β-casein (β-CN), as determined by sodium dodecyl sulfate-polyacrylamide gels (SDS-PAGE), whereas the hydrolysis of αs-casein (αs-CN) was slow and comparable to that caused by chymosin and asparatic acid proteinase from Rhizomucor miehei. The cleavage site of the metalloproteinase in κ-CN was located at the Met106–Ala107 bond, as determined by mass spectrometry analysis.

High Milk-Clotting Activity Expressed by the Newly Isolated Paenibacillus spp. Strain BD3526.

Paenibacillus spp. BD3526, a bacterium exhibiting a protein hydrolysis circle surrounded with an obvious precipitation zone on skim milk agar, was isolated from raw yak (Bos grunniens) milk collected in Tibet, China. Phylogenetic analysis based on 16S rRNA and whole genome sequence comparison indicated the isolate belong to the genus Paenibacillus. The strain BD3526 demonstrated strong ability to produce protease with milk clotting activity (MCA) in wheat bran broth. The protease with MCA was predominantly accumulated during the late-exponential phase of growth. The proteolytic activity (PA) of the BD3526 protease was 1.33-fold higher than that of the commercial R. miehei coagulant. A maximum MCA (6470 ± 281 SU mL−1) of the strain BD3526 was reached under optimal cultivation conditions. The protease with MCA was precipitated from the cultivated supernatant of wheat bran broth with ammonium sulfate and purified by anion-exchange chromatography. The molecular weight of the protease with MCA was determined as 35 kDa by sodium dodecyl sulfate-polyacrylamide gels electrophoresis (SDS-PAGE) and gelatin zymography. The cleavage site of the BD3526 protease with MCA in κ-casein was located at the Met106–Ala107 bond, as determined by mass spectrometry analysis.

Recent advances in exopolysaccharides from Paenibacillus spp.: production, isolation, structure, and bioactivities.

This review provides a comprehensive summary of the most recent developments of various aspects (i.e., production, purification, structure, and bioactivity) of the exopolysaccharides (EPSs) from Paenibacillus spp. For the production, in particular, squid pen waste was first utilized successfully to produce a high yield of inexpensive EPSs from Paenibacillus sp. TKU023 and P. macerans TKU029. In addition, this technology for EPS production is prevailing because it is more environmentally friendly. The Paenibacillus spp. EPSs reported from various references constitute a structurally diverse class of biological macromolecules with different applications in the broad fields of pharmacy, cosmetics and bioremediation. The EPS produced by P. macerans TKU029 can increase in vivo skin hydration and may be a new source of natural moisturizers with potential value in cosmetics. However, the relationships between the structures and activities of these EPSs in many studies are not well established. The contents and data in this review will serve as useful references for further investigation, production, structure and application of Paenibacillus spp. EPSs in various fields.