Cold Shock Proteins Promote Nisin Tolerance in Listeria monocytogenes Through Modulation of Cell Envelope Modification Responses

Listeria monocytogenes continues to be a food safety challenge owing to its stress tolerance and virulence traits. Several listeriosis outbreaks have been linked to the consumption of contaminated ready-to-eat food products. Numerous interventions, including nisin application, are presently employed to mitigate against L. monocytogenes risk in food products. In response, L. monocytogenes deploys several defense mechanisms, reducing nisin efficacy, that are not yet fully understood. Cold shock proteins (Csps) are small, highly conserved nucleic acid-binding proteins involved in several gene regulatory processes to mediate various stress responses in bacteria. L. monocytogenes possesses three csp gene paralogs; cspA, cspB, and cspD. Using a panel of single, double, and triple csp gene deletion mutants, the role of Csps in L. monocytogenes nisin tolerance was examined, demonstrating their importance in nisin stress responses of this bacterium. Without csp genes, a L. monocytogenes ΔcspABD mutant displayed severely compromised growth under nisin stress. Characterizing single (ΔcspA, ΔcspB, and ΔcspD) and double (ΔcspBD, ΔcspAD, and ΔcspAB) csp gene deletion mutants revealed a hierarchy (cspD > cspB > cspA) of importance in csp gene contributions toward the L. monocytogenes nisin tolerance phenotype. Individual eliminations of either cspA or cspB improved the nisin stress tolerance phenotype, suggesting that their expression has a curbing effect on the expression of nisin resistance functions through CspD. Gene expression analysis revealed that Csp deficiency altered the expression of DltA, MprF, and penicillin-binding protein-encoding genes. Furthermore, the ΔcspABD mutation induced an overall more electronegative cell surface, enhancing sensitivity to nisin and other cationic antimicrobials as well as the quaternary ammonium compound disinfectant benzalkonium chloride. These observations demonstrate that the molecular functions of Csps regulate systems important for enabling the constitution and maintenance of an optimal composed cell envelope that protects against cell-envelope-targeting stressors, including nisin. Overall, our data show an important contribution of Csps for L. monocytogenes stress protection in food environments where antimicrobial peptides are used. Such knowledge can be harnessed in the development of better L. monocytogenes control strategies. Furthermore, the potential that Csps have in inducing cross-protection must be considered when combining hurdle techniques or using them in a series

Levels of Household Food Insecurity in Rural Kenya: A Case Study of Kitui West, Lower Yatta and Matinyani Districts

As a general rule, food security is built on three pillars which are Availability, Accessibility and utilization. The ability to ensure adequate food security hinges on the ability to identify vulnerable households. The degree of vulnerability of an individual, household or group of persons is determined by their exposure to the risk factors and their ability to cope with or withstand stressful situations. This study was a component of a project whose aim was to transfer technology and scale up water harvesting and greenhouse farming in the area with one of its objectives to establish the levels of household food insecurity.  Using a proportionate sample of 381 households two major tools were used to collect data. These were a structured interview schedule which was used to collect demographic information and a modified Household Food Insecurity Access Scale (HFIAS) was used to collect information on household food insecurity.  In the study it was established that 60.3 percent of the households were getting their food mainly from the farms. Only 39.4 percent relied on the market as their main source of food and 0.3 depended on donations. The households in Kitui had a HFIAS score of 6.86 and falling in the class of severely food insecure with hunger. Keywords: Food Security, Food Insecurity.DOI: 10.7176/JEES/9-5-06Publication date: May 31st 201

Comparative genomics of dairy-associated Staphylococcus aureus from selected sub-Saharan African regions reveals milk as reservoir for human-and animal-derived strains and identifies a putative animal-related clade with presumptive novel siderophore

Staphylococcus aureus infection is considered to be a neglected tropical disease with huge impact on human and animal health alike. Dairy production in sub-Saharan Africa (SSA) relies heavily on various animals such as cows, goats, and camels, depending on the region. S. aureus causes mastitis and exhibits high prevalence in raw milk. The population structure including genotypic and phenotypic traits of dairy S. aureus in relation to animal and human isolates is, however, unknown for SSA. In this work, 20 S. aureus dairy isolates from East and West Africa were selected for comparative genomics and phenotypic analysis. Comparing their population structure revealed a large diversity of different origins suggesting milk to be a reservoir for human and animal strains alike. Furthermore, a novel putative siderophore was detected in multiple strains in a distinct animal-clade with strains of global origin. This putative siderophore shares a high genetic identity with that from Streptococcus equi suggesting possible horizontal gene transfer. These findings combined with the virulence genes harbored by these dairy-derived strains such as pvl, human evasion factor scn, various enterotoxin, leucocidin and antibiotic resistance genes, stresses the need for an integrative One Health approach to tackle the problem of S. aureus infections in animals and humans in sub-Saharan Africa

Unraveling the Genotypic and Phenotypic Diversity of the Psychrophilic Clostridium estertheticum Complex, a Meat Spoilage Agent

The spoilage of vacuum-packed meat by Clostridium estertheticum complex (CEC), which is accompanied by or without production of copious amounts of gas, has been linked to the acetone–butyrate–ethanol fermentation, but the mechanism behind the variable gas production has not been fully elucidated. The reconstruction and comparison of intra- and interspecies metabolic pathways linked to meat spoilage at the genomic level can unravel the genetic basis for the variable phenotype. However, this is hindered by unavailability of CEC genomes, which in addition, has hampered the determination of genetic diversity and its drivers within CEC. Therefore, the current study aimed at determining the diversity of CEC through comprehensive comparative genomics. Fifty CEC genomes from 11 CEC species were compared. Recombination and gene gain/loss events were identified as important sources of natural variation within CEC, with the latter being pronounced in genomospecies2 that has lost genes related to flagellar assembly and signaling. Pan-genome analysis revealed variations in carbohydrate metabolic and hydrogenases genes within the complex. Variable inter- and intraspecies gas production in meat by C. estertheticum and Clostridium tagluense were associated with the distribution of the [NiFe]-hydrogenase hyp gene cluster whose absence or presence was associated with occurrence or lack of pack distention, respectively. Through comparative genomics, we have shown CEC species exhibit high genetic diversity that can be partly attributed to recombination and gene gain/loss events. We have also shown genetic basis for variable gas production in meat can be attributed to the presence/absence of the hyp gene cluster

Prevalence of Os Acromiale in an Indigenous East African Population: A Computerized Tomography Scan-based Study

Background: Failure of fusion to any of the acromion physes at maturity may lead to an os acromiale. A radiological review into the prevalence of os acromiale in an indigenous East African population was performed using computer tomography (CT) imaging. The study aimed to demonstrate a lower prevalence of os acromiale in an urban population compared with cadaveric studies. Methods: This was a cross-sectional study with consecutive sampling. CT scans of both shoulders obtained for non-shoulder pathology were used for the analysis. CT scanning was performed at a tertiary referral hospital from January 2019 to July 2020. Results: Nine hundred eighty-two CT studies were eligible for the review. There were 148 os acromiale identified that were either pre-acromiale or mesoacromiale. The study found a 15% prevalence of os acromiale among an ethnic East African population, of which almost 40% were meso-acromiale and 60% were pre-acromiale, with no meta-acromiale. Conclusion: The prevalence of os acromiale found in this study is similar to that observed in earlier cadaveric studies

Rain Water Harvesting for Enhanced Household Water, Food and Nutritional Security: Case Study of Kitui West, Lower Yatta and Matinyani Districts, Kenya

In semi-arid areas of Kenya, over dependence on agriculture, low and unreliable rainfall and high livestock numbers, justify the considerations of rainwater harvesting for agricultural production. A step by step is now taken by the respective government agencies and non-governmental organizations to promote rainwater harvesting, as well as promoting it to the public. This study conducted a focused evaluation of the residents’ experience towards the effectiveness of rainwater harvesting system in the context of minimizing the environmental problems, the benefits associated with rainwater harvesting, the implementation issue in relation to rainwater harvesting and adoption of greenhouse technology as a paradigm shift from open field cultivation. Research was conducted among 387 respondents: 109 in Kitui West, 104 in Matinyani and 124 in Lower Yatta districts of Kitui County, Kenya. The methodology adopted used semi-structured questionnaire survey and the data was analyzed using the SPSS. The study revealed that 56.3 % of the families are female- headed while 43.7% are male headed. Agriculture is a common practice in the study area because 90.4 % of the respondents had above ½ of their lands utilized.  However, it is practiced using traditional farm implements such as hoes and simple harvesting implements as affirmed by 46.3 % of the respondents. It was found that water pans are the main sources of water in the area with only 67.2 % able to access it. However, only 55.2 % of the households could access water of above 200 liters on a daily basis and owing to the average family size of 4-6 members in the study area. The use of greenhouse technology is not popular because only 11.3 % of the respondents owned greenhouses while 85.8 % of the respondents said they wanted to own greenhouses. Surprisingly, those who own greenhouse have never made money from this technology. Keywords: Rainwater harvesting, semi-arid areas, greenhouse technology, food security, respondent

Targeted Genome Mining Reveals the Psychrophilic Clostridium estertheticum Complex as a Potential Source for Novel Bacteriocins, Including Cesin A and Estercticin A

Antimicrobial resistance in pathogenic bacteria is considered a major public health issue necessitating the discovery of alternative antimicrobial compounds. In this regard, targeted genome mining in bacteria occupying under-explored ecological niches has the potential to reveal such compounds, including bacteriocins. In this study, we determined the bacteriocin biosynthetic potential of the psychrophilic Clostridium estertheticum complex (CEC) through a combination of genome mining and phenotypic screening assays. The genome mining was performed in 40 CEC genomes using antiSMASH. The production of bacteriocin-like compounds was phenotypically validated through agar well (primary screening) and disk diffusion (secondary screening) assays using cell free supernatants (CFS) and partially purified extracts, respectively. Stability of four selected CFS against proteolytic enzymes, temperature and pH was determined while one CFS was analyzed by HRMS and MS/MS to identify potential bacteriocins. Twenty novel bacteriocin biosynthetic gene clusters (BBGC), which were classified into eight (six lantibiotics and two sactipeptides) distinct groups, were discovered in 18 genomes belonging to C. estertheticum (n = 12), C. tagluense (n = 3) and genomospecies2 (n = 3). Primary screening linked six BBGC with narrow antimicrobial activity against closely related clostridia species. All four preselected CFS retained activity after exposure to different proteolytic, temperature and pH conditions. Secondary screening linked BBGC1 and BBGC7 encoding a lantibiotic and sactipeptide, respectively, with activity against Bacillus cereus while lantibiotic-encoding BBGC2 and BBGC3 were linked with activity against B. cereus, Staphylococcus aureus (methicillin-resistant), Escherichia coli and Pseudomonas aeruginosa. MS/MS analysis revealed that C. estertheticum CF004 produces cesin A, a short natural variant of nisin, and HRMS indicated the production of a novel sactipeptide named estercticin A. Therefore, we have shown the CEC, in particular C. estertheticum, is a source of novel and stable bacteriocins that have activities against clinically relevant pathogens

Interrogating the Diversity of Vaginal, Endometrial, and Fecal Microbiomes in Healthy and Metritis Dairy Cattle

The bovine genital tract harbors a dynamic microbiome. Genital tract microbial communities in healthy animals have been characterized using next-generation sequencing methods showing that microbe compositions differ between the vagina and uterus, more so during the postpartum period. Pre-calving fecal and vaginal, and endometrial swabs at the different postpartum intervals were collected from dairy cows. Microbiomes in these samples were determined based on bacterial 16S amplicon sequencing and compared between healthy (H; n = 10) control animals and cows that developed metritis (M; n = 10) within 21 days postpartum (DPP). Compared to healthy animals the pre-calving fecal and vaginal microbiomes of metritis animals were more abundant in sequences from the phylum Fusobacteria and the bacterial genera such as Escherichia-Shigella and Histophilus. In addition, compared to healthy animals, metritis cows harboured low microbial species diversity in the endometrium, as well as decreasing Proteobacteria and increasing Fusobacteria, Firmicutes, Actinobacteria, and Bacteroidetes abundances. The greatest taxonomic compositional deviations in endometrial microbial communities between the metritis and health cows were detected between 7 and 10 DPP. There was high taxonomic similarity detected between postpartum endometrial microbiomes and the prepartum vaginal and fecal microbiomes suggesting that colonization through bacteria ascending from the rectum and vagina to the uterine cavity might play a major role in establishing the endometrial microbiome postpartum. A deeper understanding of the establishment and dynamics of postpartum endometrial microbial communities in cows will thus provide crucial basic knowledge to guide the development of genital microbiome manipulation strategies for preventing uterine disease and improving fertility in dairy cows

Draft Genome Sequences of Enterococcus mundtii Strains Isolated from Beef Slaughterhouses in Kenya

We present here draft genome sequences of Enterococcus mundtii strains K7-EM, P2-EM, C11-EM, and H18-EM, which were isolated from slaughterhouse equipment, carcasses, and personnel of small- and medium-sized beef slaughterhouses in Kenya

Species distribution and antimicrobial profiles of Enterococcus spp. isolates from Kenyan small and medium enterprise slaughterhouses

The present study aimed at identifying and assessing antimicrobial resistance of Enterococcus spp. isolated from small and medium enterprise slaughterhouses in Kenya. In total, 67 isolates were recovered from 48 of 195 samples examined from beef carcasses, personnel, and cutting equipment in five small and medium enterprise slaughterhouses. The isolates were identified by using matrix-assisted laser desorption-ionization time of flight mass spectrometry and screened thereafter for their resistance against 12 antibiotics by using a disk diffusion assay. The isolates ( n = 67) included Enterococcus faecalis (41.8%), Enterococcus mundtii (17.9%), Enterococcus thailandicus (13.4%), Enterococcus faecium (9.0%), Enterococcus hirae (7.5%), Enterococcus casseliflavus (6.0%), and Enterococcus devriesei (4.5%). None of the isolates were resistant to ciprofloxacin, penicillin, ampicillin, vancomycin, nitrofurantoin, teicoplanin, linezolid, and levofloxacin. Resistance to rifampin (46.3%), erythromycin (23.9%), tetracycline (20.9%), and chloramphenicol (7.5%) was distributed among six of the seven species. All E. thailandicus were resistant to rifampin, erythromycin, and tetracycline. E. faecalis was resistant to rifampin (60.7%), tetracycline (17.9%), erythromycin (14.3%), and chloramphenicol (10.7%). Resistance to two or three antibiotics was observed in 26.9% of the enterococci isolates. The isolation of enterococci that are resistant to clinically relevant antibiotics, such as erythromycin, is of a serious concern given the role enterococci play in the transfer of antibiotic resistance genes