Camel Milk Resistome in Kuwait: Genotypic and Phenotypic Characterization.

Antimicrobial resistance (AMR) is one of the major global health and economic threats. There is growing concern about the emergence of AMR in food and the possibility of transmission of microorganisms possessing antibiotic resistance genes (ARGs) to the human gut microbiome. Shotgun sequencing and in vitro antimicrobial susceptibility testing were used in this study to provide a detailed characterization of the antibiotic resistance profile of bacteria and their ARGs in dromedary camel milk. Eight pooled camel milk samples, representative of multiple camels distributed in the Kuwait desert, were collected from retail stores and analyzed. The genotypic analysis showed the presence of ARGs that mediate resistance to 18 classes of antibiotics in camel milk, with the highest resistance to fluoroquinolones (12.48%) and disinfecting agents and antiseptics (9%). Furthermore, the results pointed out the possible transmission of the ARGs to other bacteria through mobile genetic elements. The in vitro antimicrobial susceptibility testing indicated that 80% of the isolates were resistant to different classes of antibiotics, with the highest resistance observed against three antibiotic classes: penicillin, tetracyclines, and carbapenems. Multidrug-resistant pathogens including Klebsiella pneumoniae, Escherichia coli, and Enterobacter hormaechei were also revealed. These findings emphasize the human health risks related to the handling and consumption of raw camel milk and highlight the necessity of improving the hygienic practices of farms and retail stores to control the prevalence of ARGs and their transmission.

Transcriptome analysis of Sparidentex hasta larvae exposed to water-accommodated fraction of Kuwait crude oil.

Anthropogenic activities have been shown to significantly affect marine life. Water pollution and oil spills are particularly deleterious to the fish population, especially during their larval stage. In this study, Sobaity-sea bream Sparidentex hasta (Valenciennes, 1830) larvae were exposed to serial dilutions of water-accommodated fraction of Kuwait crude oil (KCO-WAF) for varying durations (3, 6, 24, 48, 72 or 96 h) in acute exposure regime. Gene expression was assessed using RNA sequencing and validated through RT-qPCR. The RNA sequencing data were aligned to the sequenced genome, and differentially expressed genes were identified in response to treatment with or without KCO-WAF at various exposure times. The highest number of differentially expressed genes was observed at the early time point of 6 h of post-exposure to KCO-WAF. The lowest number of differentially expressed genes were noticed at 96 h of treatment indicating early response of the larvae to KCO-WAF contaminant. The acquired information on the differentially expressed genes was then used for functional and pathway analysis. More than 90% of the differentially expressed genes had a significant BLAST match, with the two most common matching species being Acanthopagrus latus and Sparus aurata. Approximately 65% of the differentially expressed genes had Gene Ontology annotations, whereas > 35% of the genes had KEGG pathway annotations. The differentially expressed genes were found to be enriched for various signaling pathways (e.g., MAPK, cAMP, PI3K-Akt) and nervous system-related pathways (e.g., neurodegeneration, axon guidance, glutamatergic synapse, GABAergic synapse). Early exposure modulated the signaling pathways, while KCO-WAF exposure of larvae for a longer duration affected the neurodegenerative/nervous system-related pathways. RT-qPCR analysis confirmed the differential expression of genes at each time point. These findings provide insights into the underlying molecular mechanisms of the deleterious effects of acute exposure to oil pollution-on marine fish populations, particularly at the early larval stage of Sparidentex hasta.

Draft genome sequence data of Enterococcus faecium R9, a multiple enterocins-producing strain.

Food contamination by pathogens results in serious health problems and economic losses. Chemical food preservatives pose a risk to human health when used in food preservation. To increase the shelf life of the products and prevent spoilage, the dairy sector is considering natural preservatives such the ribosomally synthesized peptides, bacteriocins. Here we present the draft genome sequence of Enterococcus faecium strain R9 producing three bacteriocins isolated from raw camel milk. These bacteriocins showed valuable technological properties, such as sensitivity to proteolytic enzymes, heat stability, and wide range of pH tolerance. The 2 × 250 bp paired end reads sequencing was performed on Illumina HiSeq 2500 sequencing. The genome sequence consisted of 3,598,862 bases, with a GC content of 37.94% bases. The number of raw reads was 4,670,510, and the assembly N50 score was 65,355 bp with a 310.28 average coverage. A total of 3,086 coding sequences (CDSs) was predicted with 2,126 CDSs with a known function and 127 with a signal peptide. Annotation of the genome sequence revealed bacteriocins encoding genes, namely, enterocin B, enterocin P, and two-component enterocin X (X-alfa and X-beta subunits). These enterocins are beneficial for controlling Listeria monocytogenes in the food industry. Genome sequence of Enterococcus faecium R9 has been deposited at the gene bank under BioSample accession number JALJED000000000 and are available in Mendeley Data [1].

Antibiotic Resistance Genes in Aerosols: Baseline from Kuwait.

Antimicrobial resistance (AMR) is one of the biggest threats to human health worldwide. The World Health Organization (WHO, Geneva, Switzerland) has launched the "One-Health" approach, which encourages assessment of antibiotic-resistant genes (ARGs) within environments shared by human-animals-plants-microbes to constrain and alleviate the development of AMR. Aerosols as a medium to disseminate ARGs, have received minimal attention. In the present study, we investigated the distribution and abundance of ARGs in indoor and outdoor aerosols collected from an urban location in Kuwait and the interior of three hospitals. The high throughput quantitative polymerase chain reaction (HT-qPCR) approach was used for this purpose. The results demonstrate the presence of aminoglycoside, beta-lactam, fluoroquinolone, tetracycline, macrolide-lincosamide-streptogramin B (MLSB), multidrug-resistant (MDR) and vancomycin-resistant genes in the aerosols. The most dominant drug class was beta-lactam and the genes were IMP-2-group (0.85), Per-2 group (0.65), OXA-54 (0.57), QnrS (0.50) and OXA-55 (0.55) in the urban non-clinical settings. The indoor aerosols possessed a richer diversity (Observed, Chao1, Shannon's and Pielou's evenness) of ARGs compared to the outdoors. Seasonal variations (autumn vs. winter) in relative abundances and types of ARGs were also recorded (R2 of 0.132 at p < 0.08). The presence of ARGs was found in both the inhalable (2.1 µm, 1.1 µm, 0.7 µm and < 0.3 µm) and respirable (>9.0 µm, 5.8 µm, 4.7 µm and 3.3 µm) size fractions within hospital aerosols. All the ARGs are of pathogenic bacterial origin and are hosted by pathogenic forms. The findings present baseline data and underpin the need for detailed investigations looking at aerosol as a vehicle for ARG dissemination among human and non-human terrestrial biota.

Genome survey and genetic characterization of Acacia pachyceras O. Schwartz.

Acacia pachyceras O. Schwartz (Leguminoseae), a woody tree growing in Kuwait is critically endangered. High throughput genomic research is immediately needed to formulate effective conservation strategies for its rehabilitation. We therefore, performed a genome survey analysis of the species. Whole genome sequencing generated ~97 Gb of raw reads (92x coverage) with a per base quality score above Q30. The k-mer analysis (17 mer) revealed its genome to be 720Mb in size with an average guanine-cytosine (GC) ratio of 35%. The assembled genome was analyzed for repeat regions (45.4%-interspersed repeats; 9%-retroelements; 2%-DNA transposons). BUSCO assessment of completeness of genome identified 93% of assembly to be complete. Gene alignments in BRAKER2 yielded 34,374 transcripts corresponding to 33,650 genes. Average length of coding sequences and protein sequences were recorded as 1,027nts and 342aa, respectively. GMATA software filtered a total of 901,755 simple sequence repeats (SSRs) regions against which 11,181 unique primers were designed. A subset of 110 SSR primers were PCR validated and demonstrated for its application in genetic diversity analysis of Acacia. The SSR primers successfully amplified A. gerrardii seedlings DNA depicting cross transferability among species. The principal coordinate analysis and the split decomposition tree (bootstrapping runs of 1000 replicates) distributed the Acacia genotypes into two clusters. The flow cytometry analysis revealed the A. pachyceras genome to be polyploid (6x). The DNA content was predicted as 2.46 pg, 1.23 pg, and 0.41 pg corresponding to 2C DNA, 1C DNA and 1Cx DNA, respectively. The results provide a base for further high throughput genomic studies and molecular breeding for its conservation.

Metagenomes from Coastal Sediments of Kuwait: Insights into the Microbiome, Metabolic Functions and Resistome.

Coastal sediments in the proximity of wastewater and emergency outfalls are often sinks of pharmaceutical compounds and other organic and inorganic contaminants that are likely to affect the microbial community. The metabolites of these contaminants affect microbial diversity and their metabolic processes, resulting in undesirable effects on ecosystem functioning, thus necessitating the need to understand their composition and functions. In the present investigation, we studied the metagenomes of 12 coastal surface sediments through whole genome shot-gun sequencing. Taxonomic binning of the genes predicted about 86% as bacteria, 1% as archaea, >0.001% as viruses and Eukaryota, and 12% as other communities. The dominant bacterial, archaeal, and fungal genera were Woeseia, Nitrosopumilus, and Rhizophagus, respectively. The most prevalent viral families were Myoviridae and Siphoviridae, and the T4 virus was the most dominant bacteriophage. The unigenes further aligned to 26 clusters of orthologous genes (COGs) and five carbohydrate-active enzymes (CAZy) classes. Glycoside hydrolases (GH) and glycoside transferase (GT) were the highest-recorded CAzymes. The Kyoto Encyclopedia of Genes and Genomes (KEGG) level 3 functions were subjugated by purine metabolism > ABC transporters > oxidative phosphorylation > two-component system > pyrimidine metabolism > pyruvate metabolism > quorum sensing > carbon fixation pathways > ribosomes > and glyoxalate and dicarboxylate metabolism. Sequences allying with plasmids, integrons, insertion sequences and antibiotic-resistance genes were also observed. Both the taxonomies and functional abundances exhibited variation in relative abundances, with limited spatial variability (ANOVA p > 0.05; ANOSIM-0.05, p > 0.05). This study underlines the dominant microbial communities and functional genes in the marine sediments of Kuwait as a baseline for future biomonitoring programs.

Assessment of mastitis in camel using high-throughput sequencing.

Camel milk is recognized as a functional food with significant economic value. Mastitis is one of the most common and costly diseases in the dairy industry. Mastitis, which is caused by pathogens such as bacteria, viruses, fungi, and algae, has an impact on the quality and quantity of milk produced as well as animal health and welfare. There is a paucity of data on the etiological factors that cause camel mastitis. This study reports the bacterial and fungal community involved in clinical camel mastitis using Illumina amplicon sequencing. A total of 25 milk samples were analyzed, including 9 samples with mastitis and 16 healthy samples. The bacterial community in healthy samples was significantly more diverse and abundant than in mastitis samples. The fungal population in mastitis samples, on the other hand, was more diverse and abundant. As compared to healthy samples, the genera Staphylococcus, Streptococcus, Schlegelella, unclassified Enterobacteriaceae, Lactococcus, Jeotgalicoccus. and Klebsiella were found to be abundant in mastitic milk. However, the genera Corynebacterium, Enteractinococcus, unclassified Sphingomonadaceae, Atopostipes, Paenibacillus, Pseudomonas, Lactobacillus, Sphingomonas, Pediococcus and Moraxella were reduced. In the fungal community, mastitis caused a significant increase in the relative abundance of the majority of taxa, including Candida, Phanerochaete, Aspergillus, Cladosporium and unclassified Pyronemataceae, while Penicillium and Alternaria showed a decline in relative abundance. In the bacterial and fungal communities, the discriminant analysis showed 19 and 5 differently abundant genera in healthy milk and mastitic milk, respectively. In conclusion, this study showed a microbiome dysbiosis linked to clinical camel mastitis, with opportunistic pathogens outgrowing commensal bacteria that were reduced. These findings are essential in designing an appropriate control program in the camel dairy herd, as well as in preventing and treating camel mastitis.

De-novo transcriptome assembly and analysis of lettuce plants grown under red, blue or white light.

Lettuce (Lactuca sativa) is grown in various parts of the world for use as a leafy vegetable. Although the use of light-emitting diode (LED) in controlled plant production systems has been successfully used to enhance nutritional quality and plant growth efficiently, the molecular basis of lettuce's response to varying light spectra is not studied. Using next-generation sequencing, we have analyzed the transcriptomes of leaf lettuce (Lactuca sativa var. 'New Red Fire') grown hydroponically in a modular agricultural production system under three different types of LED lighting: red, blue, and white light. Illumina HiSeq sequencing platform was used to generate paired-end sequence reads (58 Gb raw and 54 Gb clean data) of the transcriptome of lettuce leaves exposed to varying light spectra. The de novo assembled final transcriptome contained 74,096 transcripts. Around 53% and 39% of the assembled transcripts matched to the UniProt and RefSeq RNA sequences, respectively. The validation of the differentially expressed transcripts using RT-qPCR showed complete agreement with RNA-Seq data for 27 transcripts. A comparison of the blue versus red light treatments showed the highest number of significantly differentially expressed transcripts. Among the transcripts significantly up-regulated in blue-light-exposed leaves compared to white-light-exposed leaves, ~ 26% were involved in the 'response to stress'. Among the transcripts significantly upregulated under red light compared to white light, ~ 6% were associated with 'nucleosome assembly' and other processes, such as 'oxidation-reduction process' and 'response to water deprivation' were significantly enriched. Thus, the result from the current study provides deeper insights into differential gene expression patterns and associated functional aspects under varying light qualities.

De-novo genome assembly and annotation of sobaity seabream Sparidentex hasta.

Sparidentex hasta (Valenciennes, 1830) of the Sparidae family, is an economically important fish species. However, the genomic studies on S. hasta are limited due to the absence of its complete genome. The goal of the current study was to sequence, assemble, and annotate the genome of S. hasta that will fuel further research related to this seabream. The assembled draft genome of S. hasta was 686 Mb with an N50 of 80 Kb. The draft genome contained approximately 22% repeats, and 41,201 genes coding for 44,555 transcripts. Furthermore, the assessment of the assembly completeness was estimated based on the detection of ∼93% BUSCOs at the protein level and alignment of >99% of the filtered reads to the assembled genome. Around 68% of the predicted proteins (n = 30,545) had significant BLAST matches, and 30,473 and 13,244 sequences were mapped to Gene Ontology annotations and different enzyme classes, respectively. The comparative genomics analysis indicated S. hasta to be closely related to Acanthopagrus latus. The current assembly provides a solid foundation for future population and conservation studies of S. hasta as well as for investigations of environmental adaptation in Sparidae family of fishes. Value of the Data: This draft genome of S. hasta would be very applicable for molecular characterization, gene expression studies, and to address various problems associated with pathogen-associated immune response, climate adaptability, and comparative genomics. The accessibility of the draft genome sequence would be useful in understanding the pathways and functions at the molecular level, which may further help in improving the economic value and their conservation.

Data on microbial diversity of camel milk microbiota determined by 16S rRNA gene sequencing.

Raw camel milk samples were collected from three geographical locations (south, north and middle Kuwait) during two seasons. Next generation sequencing of the V3-V4 regions of the 16S rRNA gene was used to analyze the bacterial community in camel milk. DNA was extracted from one hundred thirty-three samples, and libraries were prepared using custom fusion primers of the 16S rRNA gene and sequenced on Illumina HiSeq 2500 platform. 16S rRNA gene sequences were aligned against the SILVA database SSU release 138. The high-throughput sequencing data are available at the NCBI database under the Bioproject PRJNA814013. This work describes camel milk's bacterial diversity among different geographical locations and seasons. The distribution of alpha diversity measures among camel milk sample groups collected from different geographical locations and seasons is presented. A significant effect of these parameters on camel milk's bacterial diversity was shown. Linear discriminant analysis (LefSe) showed significant differentially abundant bacteria at the phylum, class, order, family and genus level among the three locations and seasons. LefSe identified a total of 83 and 40 differentially abundant genera in the different geographical locations and seasons, respectively. More details about the bacterial composition of raw camel milk at the phylum and genus level can be found in research article [1]. These data can be used to compare the diversity of milk bacterial community between different milk producing species and camels from different parts of the world. Besides, these findings will contribute to our understanding of the camel microbiome structure and might be useful for designing an appropriate control program in the camel dairy herd. The data described in this article are available in Mendeley Data [2].

Bacterial and fungal communities in indoor aerosols from two Kuwaiti hospitals.

The airborne transmission of COVID-19 has drawn immense attention to bioaerosols. The topic is highly relevant in the indoor hospital environment where vulnerable patients are treated and healthcare workers are exposed to various pathogenic and non-pathogenic microbes. Knowledge of the microbial communities in such settings will enable precautionary measures to prevent any hospital-mediated outbreak and better assess occupational exposure of the healthcare workers. This study presents a baseline of the bacterial and fungal population of two major hospitals in Kuwait dealing with COVID patients, and in a non-hospital setting through targeted amplicon sequencing. The predominant bacteria of bioaerosols were Variovorax (9.44%), Parvibaculum (8.27%), Pseudonocardia (8.04%), Taonella (5.74%), Arthrospira (4.58%), Comamonas (3.84%), Methylibium (3.13%), Sphingobium (4.46%), Zoogloea (2.20%), and Sphingopyxis (2.56%). ESKAPEE pathogens, such as Pseudomonas, Acinetobacter, Staphylococcus, Enterococcus, and Escherichia, were also found in lower abundances. The fungi were represented by Wilcoxinia rehmii (64.38%), Aspergillus ruber (9.11%), Penicillium desertorum (3.89%), Leptobacillium leptobactrum (3.20%), Humicola grisea (2.99%), Ganoderma sichuanense (1.42%), Malassezia restricta (0.74%), Heterophoma sylvatica (0.49%), Fusarium proliferatum (0.46%), and Saccharomyces cerevisiae (0.23%). Some common and unique operational taxonomic units (OTUs) of bacteria and fungi were also recorded at each site; this inter-site variability shows that exhaled air can be a source of this variation. The alpha-diversity indices suggested variance in species richness and abundance in hospitals than in non-hospital sites. The community structure of bacteria varied spatially (ANOSIM r 2 = 0.181-0.243; p < 0.05) between the hospital and non-hospital sites, whereas fungi were more or less homogenous. Key taxa specific to the hospitals were Defluvicoccales, fungi, Ganodermataceae, Heterophoma, and H. sylvatica compared to Actinobacteria, Leptobacillium, L. leptobacillium, and Cordycipitaceae at the non-hospital site (LefSe, FDR q ≤ 0.05). The hospital/non-hospital MD index > 1 indicated shifts in the microbial communities of indoor air in hospitals. These findings highlight the need for regular surveillance of indoor hospital environments to prevent future outbreaks.

Comparison and optimization of DNA Isolation protocols for high throughput genomic studies of Acacia pachyceras Schwartz.

We describe the optimization and validation of six DNA isolation protocols from fresh leaves of the rare tree Acacia pachyceras. The first four protocols employed three commercial kits (Sigma, Nucleospin1, Nucleospin 2, Promega) whereas the remaining two were based on the traditional sodium dodecyl sulfate (SDS) and cetyltrimethylammonium bromide CTAB methods. Each protocol provided significantly different results concerning DNA concentration (p < 0.032), yield (p < 0.000), contaminant carry over, protocol duration, cost per sample, and comprehensive cost. We demonstrated the applicability of all the tested protocols in DNA barcoding. The protocol yielded maximum amounts (92.85 µg) of DNA in a rapid turnaround time (8 h). The quantity and purity surpassed all the other tested methods. DNA extracted by the CTAB method was the best for NGS (Phred score >Q30). These protocols will be useful tools for molecular research of Acacia pachyceras and other closely related tree species.

Camel milk microbiota: A culture-independent assessment.

Camel milk is renowned for its nutritional value and its therapeutic properties. It is considered a promising alternative to bovine milk due to its higher nutritional benefits, hypoallergenic characteristics and greater digestibility in the human gastrointestinal system. This study reports camel milk's bacterial and fungal microbiota, and the effect of geographical location and season on its bacterial community. We sequenced the V3-V4 regions of the16S rRNA gene for bacteria and the internal transcribed spacer (ITS) for fungi. A total of 134 samples of dromedary raw camel milk were collected from south, north and middle Kuwait during two seasons. Raw camel milk showed a diversified bacterial community, with 1196 genera belonging to 33 phyla. The four most predominant phyla of bacteria were Proteobacteria, Firmicutes, Actinobacteria and Bacteroidota. The core microbiota of raw camel milk, represented by the dominant genera shared by the majority of samples, was constituted by the genera Schlegelella, Paenibacillus, Lactobacillus, unclassified Comamonadaceae, Pediococcus, Moraxella, Acinetobacter, Staphylococcus, Enterococcus, Pseudomonas, Streptococcus, unclassified Micrococcaceae, Rothia, unclassified Sphingomonadaceae, unclassified Neisseriaceae and Sphingomonas. The fungal population was assessed in 14 raw camel milk samples, and comprised 87 genera belonging to 3 phyla. The genera Penicillium, Cladosporium, Candida, Aspergillus, Alternaria and Fusarium, dominated the fungal community. These findings shed light on raw camel milk's core bacterial and fungal microbiome. The geographical location and the season had a significant impact on the diversity and composition of camel milk microbiome.

Draft genome sequence and SSR mining data of Acacia pachyceras Schwartz.

Acacia tree population is declining in several countries of the world especially in the Arabian peninsula due to human-induced activities. The tree has potential medicinal and economic benefits as a source of fuel and timber. It can fix nitrogen, a significant property that assists in desert rehabilitation. However, the lack of genomic information of Acacia pachyceras hampers its genetic study and breeding process. We performed paired-end sequencing of A. pachyceras at a depth of 120X to obtain raw sequences of 108.9 GB with a per base quality >Q30. Filtered raw data was assembled into a fasta file of 4 GB. The assembled genomic sequences consisted of 901,755 single sequence repeats (SSRs). In total 11,596 primer pairs were designed against these SSR motifs. The data generated provides baseline genomic information about the species and formulates a base for further sequencing of A. pachyceras through PACBio and HiC technologies. The novel developed SSR markers will facilitate genetic diversity and conservation studies for Acacia species.

SARS-CoV-2 in hospital air as revealed by comprehensive respiratory viral panel sequencing.

BACKGROUND: Nosocomially acquired severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) infection has become the most significant pandemic of our lifetime. Though its transmission was essentially attributed to droplets from an infected person, with recent advancements in knowledge, aerosol transmission seems to be a viable pathway, as well. Because of the lower biological load in ambient aerosol, detection of SARS-CoV-2 is challenging. A few recent attempts of sampling large aerosol volumes and using next-generation sequencing (NGS) to detect the presence of SARS-CoV-2 in the air at very low levels gave positive results. These results suggest the potential of using this technique to detect the presence of SARS-CoV-2 and use it as an early warning signal for possible outbreak or recurrence of coronavirus disease 2019 (COVID-19). AIM: To assess efficacy of comprehensive respiratory viral panel (CRVP) sequencing and RT-PCR for low-level identification of SARS-CoV-2 and other respiratory viruses in indoor air. METHODS: A large volume of indoor aerosol samples from three major hospitals involved in COVID-19 care in Kuwait was collected. Viral RNA was isolated and subjected to comprehensive respiratory viral panel sequencing (CRVP) as per the standard protocol to detect the SARS-CoV-2 and other respiratory viruses in the hospital aerosol and monitor variations within the sequences. RT-PCR was also employed to estimate the viral load of SARS-CoV-2. FINDINGS: 13 of 15 (86.7%) samples exhibited SARS-CoV-2 with a relative abundance of 0.2-33.3%. The co-occurrence of human adenoviruses (type C1, C2, C5, C4), respiratory syncytial virus (RSV), influenza B, and non-SARS-CoV-229E were also recorded. Alignment of SARS-CoV-2 sequences against the reference strain of Wuhan China revealed variations in the form of single nucleotide polymorphisms (SNPs-17), insertions and deletions (indels-1). These variations were predicted to create missense (16), synonymous (15), frameshift (1) and stop-gained (1) mutations with a high (2), low (15), and moderate (16) impact. CONCLUSIONS: Our results suggest that using CRVP on a large volume aerosol sample was a valuable tool for detecting SARS-CoV-2 in indoor aerosols of health care settings. Owing to its higher sensitivity, it can be employed as a surveillance strategy in the post COVID times to act as an early warning system to possibly control future outbreaks.

Insights into Bacterial Community Involved in Bioremediation of Aged Oil-Contaminated Soil in Arid Environment.

Soil contamination by hydrocarbons due to oil spills has become a global concern and it has more implications in oil producing regions. Biostimulation is considered as one of the promising remediation techniques that can be adopted to enhance the rate of degradation of crude oil. The soil microbial consortia play a critical role in governing the biodegradation of total petroleum hydrocarbons (TPHs), in particular polycyclic aromatic hydrocarbons (PAHs). In this study, the degradation pattern of TPHs and PAHs of Kuwait soil biopiles was measured at three-month intervals. Then, the microbial consortium associated with oil degradation at each interval was revealed through 16S rRNA based next generation sequencing. Rapid degradation of TPHs and most of the PAHs was noticed at the first 3 months of biostimulation with a degradation rate of pyrene significantly higher compared to other PAHs counterparts. The taxonomic profiling of individual stages of remediation revealed that, biostimulation of the investigated soil favored the growth of Proteobacteria, Alphaprotobacteria, Chloroflexi, Chlorobi, and Acidobacteria groups. These findings provide a key step towards the restoration of oil-contaminated lands in the arid environment.

Distribution and diversity of eukaryotic microalgae in Kuwait waters assessed using 18S rRNA gene sequencing.

The microbial communities play a crucial role in ecosystem functioning through interactions among individuals and taxonomic groups in a highly dynamic marine ecosystem. The structure and functioning of the microbial communities are often influenced by the changes in the surrounding environment. Monitoring the microbial diversity of the marine ecosystem helps to understand spatial patterns of microbial community and changes due to season, climate, and various drivers of biological diversity. Kuwait is characterized by an arid environment with a high degree of temperature variation during summer and winter. Our understanding of spatial distribution patterns of microbial communities, their diversity, and the influence of human activities on the degree of changes in the diversity of the microbial community in Kuwait territorial waters remain unclear. In this study, we employed 18S rRNA sequencing to explore marine microalgal community composition and dynamics in seawater samples collected from Kuwait waters over two seasonal cycles across six locations. A total of 448,184 sequences across 36 replicates corresponding to 12 samples from six stations were obtained. The quality-filtered sequences were clustered into 1,293 representative sequences, which were then classified into different eukaryotic taxa. This study reveals that the phytoplankton community in Kuwait waters is diverse and shows significant variations among different taxa during summer and winter. Dinoflagellates and diatoms were the most abundant season-dependent microalgae taxa in Kuwait waters. Alexandrium and Pyrophacus were abundant in summer, whereas Gonyaulax was abundant during the winter. The abundance of Coscinodiscus and Navicula, of the diatom genera, were also dependent upon both seasonal and possible anthropogenic factors. Our results demonstrate the effectiveness of a sequencing-based approach, which could be used to improve the accuracy of quantitative eukaryotic microbial community profiles.

Characterization and application of antimicrobials produced by Enterococcus faecium S6 isolated from raw camel milk.

The emergence of antimicrobial resistance in the food chain and the consumer's demand for safe food without chemical preservatives have generated much interest in natural antimicrobials. Thus, our main goal was to study the mode of action of the crude extract, the enterocins, and the organic acid produced by a bacteriocinogenic Enterococcus faecium strain S6 previously isolated from raw camel milk. Then, we aimed to evaluate their potential application in a food system. These antimicrobials exhibited antimicrobial activity against Listeria monocytogenes, Salmonella enterica, and Escherichia coli. The enterocins were synthesized as primary metabolites beginning at the lag phase, with optimal production at the exponential and stationary phases. The antimicrobials had a direct effect in extending the lag phase of L. monocytogenes, along with a significant inhibitory activity. The organic acid, in particular, inhibited both L. monocytogenes and S. enterica by inducing a total lysis and damage of the cell wall. The enterocins acted on disrupting the cell wall with pore formation, leading to cell death. Moreover, the crude extract revealed a combined inhibitory activity between enterocins and organic acid. Furthermore, the antimicrobials showed promising results through inhibiting L. monocytogenes cells in milk samples up to 1 wk at 4°C.

Metagenomic analysis of rhizosphere microflora of oil-contaminated soil planted with barley and alfalfa.

The role of rhizosphere microbial communities in the degradation of hydrocarbons remains poorly understood and is a field of active study. We used high throughput sequencing to explore the rhizosphere microbial diversity in the alfalfa and barley planted oil contaminated soil samples. The analysis of 16s rRNA sequences showed Proteobacteria to be the most enriched (45.9%) followed by Bacteriodetes (21.4%) and Actinobacteria (10.4%) phyla. The results also indicated differences in the microbial diversity among the oil contaminated planted soil samples. The oil contaminated planted soil samples showed a higher richness in the microbial flora when compared to that of untreated samples, as indicated by the Chao1 indices. However, the trend was different for the diversity measure, where oil contaminated barley planted soil samples showed slightly lower diversity indices. While the clustering of soil samples grouped the oil contaminated samples within and across the plant types, the clean sandy soil samples formed a separate group. The oil contaminated rhizosphere soil showed an enrichment of known oil-degrading genera, such as Alcanivorax and Aequorivita, later being specifically enriched in the contaminated soil samples planted with barley. Overall, we found a few well known oil-degrading bacterial groups to be enriched in the oil contaminated planted soil samples compared to the untreated samples. Further, phyla such as Thermi and Gemmatimonadetes showed an enrichment in the oil contaminated soil samples, indicating their potential role in hydrocarbon degradation. The findings of the current study will be useful in understanding the rhizosphere microflora responsible for oil degradation and thus can help in designing appropriate phytoremediation strategies for oil contaminated lands.

Characterization of semipurified enterocins produced by Enterococcus faecium strains isolated from raw camel milk.

Food safety has become an issue of great interest worldwide. Listeria monocytogenes is a food-borne pathogen that causes listeriosis and is difficult to control in the dairy industry. The use of lactic acid bacteria (LAB) and their antimicrobial substances against Listeria is promising in food applications. Here, we report the isolation from raw camel milk of LAB displaying antilisterial activity. Two isolates were selected for their secretion of bacteriocin(s) and identified by 16S rRNA sequencing as Enterococcus faecium S6 and R9. The produced bacteriocins were partially purified by ammonium sulfate precipitation and then biochemically characterized. Antimicrobial activity was estimated to be 6,400 and 400 AU (arbitrary units)/mL for E. faecium S6 and R9, respectively. The proteinaceous nature of the bacteriocins was confirmed via enzymatic reactions. Moreover, lipolytic and glycolytic enzymes completely inactivated the antimicrobial effect of the bacteriocins. These bacteriocins were heat-resistant and stable over a wide range of pH (2.0 to 10.0). To confirm its inactivation by lipolytic and glycolytic enzymes, the bacteriocin of E. faecium S6 was further purified by gel filtration, which suggested the existence of carbohydrate and lipid moieties. In addition, enterocin-coding genes were identified by PCR, showing DNA fragments corresponding in size to enterocins A, B, and P for E. faecium S6 and to enterocins B and P for E. faecium R9. In conclusion, these results indicate that partially purified bacteriocins from E. faecium S6 and R9 may be beneficial in controlling Listeria in the dairy industry.