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.

Genome of Tomato Yellow Leaf Curl Virus (TYLCV) Obtained from Tomato Leaves in Kuwait.

We report here the genome sequence of tomato yellow leaf curl virus (TYLCV) Wafra 19, isolated in Kuwait from symptomatic tomato plant leaves. The genome showed 98.42% identity to the AZ23-1 strain genome sequence.

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].

Genetic recombination among tomato yellow leaf curl virus isolates in commercial tomato crops in Kuwait drives emergence of virus diversity: a comparative genomic analysis.

OBJECTIVE: Whitefly-transmitted tomato yellow leaf curl virus (TYLCV) continues to be a major constraint to tomato production in Kuwait. However, very limited information is available about the population structure and genetic diversity of TYLCV infecting tomato in Kuwait. RESULTS: Whole genome sequences of 31 isolates of TYLCV, collected from commercial tomato crops grown in northern (Abdally) and southern (Al Wafra) parts of Kuwait, were deciphered. Eighteen isolates of TYLCV are identified as potential genetic recombinants. The isolates Abdally 6A and Abdally 3B reported in this study were identified to be potential recombinants. Compared to the 15 isolates from the Abdally area, and the three previously reported KISR isolates of Kuwait, six out of sixteen Al Wafra isolates showed an insertion of 19 extra nucleotides near the 5'-end. There are also four nucleotide variations before the 19-extra-nucleotides. The additional 19 nucleotides observed in nine isolates indicate that these isolates might have resulted from a single gene recombination/insertion event. Molecular phylogeny based on complete genome sequences of TYLCV isolates suggests transboundary movement of virus isolates due to geographic proximity. The information presented herein is quite useful for the comprehension of TYLCV biology, epidemiology and would aid in the management of disease in the long run.

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.

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].

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 of Bacteriocin-Encoding Enterococcus faecium Strain S6, Isolated from Camel Milk.

Enterococcus faecium strain S6 is a newly identified bacteriocin producer isolated from raw camel milk. The draft genome sequence is composed of 2,617,971 bp, with 2,407 coding genes and a G+C content of 37.99%. The genome sequence analysis provided details into the antimicrobial properties of strain S6.

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.

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.

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.

Draft genome sequence of the silver pomfret fish, Pampus argenteus.

Silver pomfret, Pampus argenteus, is a fish species from coastal waters. Despite its high commercial value, this edible fish has not been sequenced. Hence, its genetic and genomic studies have been limited. We report the first draft genome sequence of the silver pomfret obtained using a Next Generation Sequencing (NGS) technology. We assembled 38.7 Gb of nucleotides into scaffolds of 350 Mb with N50 of about 1.5 kb, using high quality paired end reads. These scaffolds represent 63.7% of the estimated silver pomfret genome length. The newly sequenced and assembled genome has 11.06% repetitive DNA regions, and this percentage is comparable to that of the tilapia genome. The genome analysis predicted 16 322 genes. About 91% of these genes showed homology with known proteins. Many gene clusters were annotated to protein and fatty-acid metabolism pathways that may be important in the context of the meat texture and immune system developmental processes. The reference genome can pave the way for the identification of many other genomic features that could improve breeding and population-management strategies, and it can also help characterize the genetic diversity of P. argenteus.

Occurrence of Pseudomonas aeruginosa in Kuwait soil.

Environmentally ubiquitous bacteria such as Pseudomonas aeruginosa evolved mechanisms to adapt and prevail under diverse conditions. In the current investigation, strains of P. aeruginosa demonstrating high rates of crude oil utilization and tolerance to high concentrations of heavy metals were found in both crude oil-contaminated and uncontaminated sites in Kuwait, and were dominant in the contaminated sites. The incidence of P. aeruginosa in tested soils implies the definitive pattern of crude oil contamination in the selection of the bacterial population in petroleum-contaminated sites in Kuwait. Surprisingly, the unculturable P. aeruginosa in different soil samples showed significant high similarity coefficients based on 16S-RFLP analyses, implying that the unculturable fraction of existing bacterial population in environmental samples is more stable and, hence, reliable for phylogenetic studies compared to the culturable bacteria.

A 12 years audit of upper gastrointestinal endoscopic procedures.

OBJECTIVE: Evaluation of upper gastrointestinal (GI) endoscopy in terms of indications, diagnostic efficacy, and diseases diagnosed. DESIGN: Retrospective, observational case series. PLACE AND DURATION OF STUDY: DHQ Teaching Hospital, Rawalpindi, from March 1990 to December 2001. SUBJECTS AND METHODS: Patients who underwent upper GI endoscopy in 12 years were included. Upper GI endoscopies were performed according to standard protocol. Endoscopic diagnoses were based on widely accepted criteria. RESULTS: Of the 8481 patients, 4935 (58.2%) were female and 3546 (41.8%) male. Mean patient age was 40.5 years. Dyspepsia (42.6%), upper GI bleed (32.8%), and evaluation of chronic liver disease (10.2%) were common indications of the procedure. An endoscopic diagnosis was possible in 82.6% patients. Varices, gastritis, duodenitis, and combined lesions were common endoscopic diagnosis. Gastritis and duodenitis were most frequent causes of upper GI bleed. We noted more gastric ulcers compared to duodenal ulcers. Females had significantly more normal endoscopies, p-value=0.02. CONCLUSION: Upper GI endoscopy is an effective procedure. Dyspepsia evaluation is commonest indication for upper GI endoscopy in our patients. Etiology of upper GI bleed, and incidence of duodenal ulcer compared to gastric ulcer in our patients are different than described in literature. Females have significantly more normal endoscopies.