Synergy in penicillin, cephalosporin, amphenicols, and aminoglycoside against MDR S. aureus isolated from Camel milk

Abstract This study investigated the synergy testing of penicillin, cephalosporin, amphenicols, and aminoglycoside in the camel milk (n=768 samples), subsequently used for isolation of MDR S. aureus targeting mecA gene. Antibiotic susceptibility of S. aureus showed >90% isolates were sensitive to ciprofloxacin and trimethoprim and resistant against oxacillin, ampicillin, and cefoxitin. Further, 50-85% of the S. aureus were sensitive to gentamicin, oxytetracycline, and chloramphenicol and resistant against cefotaxime, vancomycin, and cefixime. Minimum inhibitory concentration (MIC) of cefotaxime, (C) and ampicillin (A) in combination with gentamicin (G) was reduced by 99.34% and 70.46%, respectively, while with chloramphenicol (Ch), reduction was 57.49% and 60%, respectively. In addition, the Fractional Inhibitory Concentration Index (FICI) of G+A, Ch+C and Ch+G combinations showed synergy against 80%, 60%, and 30% of MDR S. aureus, respectively. Similarly, C+A and Ch+G displayed indifferent interaction against 70 % and 30% of isolates, respectively, while the later showed additive interaction against 10% of MDR S. aureus. Altogether, our results described effective combination of gentamicin and chloramphenicol with ampicillin and cefotaxime to combat MDR S. aureus

Modes of genetic adaptations underlying functional innovations in the rumen.

The rumen is the hallmark organ of ruminants and hosts a diverse ecosystem of microorganisms that facilitates efficient digestion of plant fibers. We analyzed 897 transcriptomes from three Cetartiodactyla lineages: ruminants, camels and cetaceans, as well as data from ruminant comparative genomics and functional assays to explore the genetic basis of rumen functional innovations. We identified genes with relatively high expression in the rumen, of which many appeared to be recruited from other tissues. These genes show functional enrichment in ketone body metabolism, regulation of microbial community, and epithelium absorption, which are the most prominent biological processes involved in rumen innovations. Several modes of genetic change underlying rumen functional innovations were uncovered, including coding mutations, genes newly evolved, and changes of regulatory elements. We validated that the key ketogenesis rate-limiting gene (HMGCS2) with five ruminant-specific mutations was under positive selection and exhibits higher synthesis activity than those of other mammals. Two newly evolved genes (LYZ1 and DEFB1) are resistant to Gram-positive bacteria and thereby may regulate microbial community equilibrium. Furthermore, we confirmed that the changes of regulatory elements accounted for the majority of rumen gene recruitment. These results greatly improve our understanding of rumen evolution and organ evo-devo in general.

Whole-genome sequencing of 128 camels across Asia reveals origin and migration of domestic Bactrian camels.

The domestic Bactrian camels were treated as one of the principal means of locomotion between the eastern and western cultures in history. However, whether they originated from East Asia or Central Asia remains elusive. To address this question, we perform whole-genome sequencing of 128 camels across Asia. The extant wild and domestic Bactrian camels show remarkable genetic divergence, as they were split from dromedaries. The wild Bactrian camels also contribute little to the ancestry of domestic ones, although they share close habitat in East Asia. Interestingly, among the domestic Bactrian camels, those from Iran exhibit the largest genetic distance and the earliest split from all others in the phylogeny, despite evident admixture between domestic Bactrian camels and dromedaries living in Central Asia. Taken together, our study support the Central Asian origin of domestic Bactrian camels, which were then immigrated eastward to Mongolia where native wild Bactrian camels inhabit.

Genetic typing of South Kazakhstan populations' dairy camels using DNA technology.

The article presents results of genetic typing of South Kazakhstan populations' dairy camels based on DNA technology using 12 camel microsatellite loci (LCA-8, LCA-19, LCA-37, LCA-56, LCA-65, LCA-66, CMS-16, YWLL-08, YWLL-29, YWLL-38, YWLL-44 and VOLP-10). The work shows the effectiveness of the used microsatellite loci in compiling the genetic profile of Kazakh Bactrian and Arvana camel populations bred in Karatau-Moiynkum and Arys-Turkestan zones of South Kazakhstan. The allele pool of the following three populations was studied using microsatellite loci: Arvana breed ('Ussenov N.' farm and 'Syzdykbekov A.' LLP) and Kazakh Bactrian breed ('Bagdat' farm). The level of genetic diversity was determined and the degree of heterozygosity of different camel populations was shown. The characteristics of the studied camel populations are given in terms of F-statistics: according to the degree of inbreeding and the level of interpopulation and intrapopulation diversity in each of the studied camel populations. For the first time in Kazakhstan, the genotyping of South Kazakhstan populations' dairy camels was carried out using DNA technology, which provides a reliable assessment of the genetic profile of highly productive individuals by microsatellite loci for development of dairy industry in different regions of the camel breeding location.

Old World camels in a modern world - a balancing act between conservation and genetic improvement.

Old World camels have served humans in cross-continental caravans, transporting people and goods, connecting different cultures and providing milk, meat, wool and draught since their domestication around 3000-6000 years ago. In a world of modern transport and fast connectivity, these beasts of burden seem to be out-dated. However, a growing demand for sustainable milk and meat production, especially in countries affected by climate change and increasing desertification, brings dromedaries (Camelus dromedarius) and Bactrian camels (Camelus bactrianus) back onstage and into the focus of animal breeders and scientists. In this review on the molecular genetics of these economically important species we give an overview about the evolutionary history, domestication and dispersal of Old World camels, whereas highlighting the need for conservation of wild two-humped camels (Camelus ferus) as an evolutionarily unique and highly endangered species. We provide cutting-edge information on the current molecular resources and on-going sequencing projects. We cannot emphasise enough the importance of balancing the need for improving camel production traits with maintaining the genetic diversity in two domestic species with specific physiological adaptation to a desert environment.

Morphometric and genetic variation in 8 breeds of Ethiopian camels (Camelus dromedarius).

Dromedary camels (Camelus dromedarius) are a domesticated and closely guarded economic staple of indigenous people located throughout Ethiopian territorial states. Seventeen morphometric variables were examined to determine intraspecific variation among 8 pastoralist-designated breeds of camels. Additionally, DNA sequences from mitochondrial cytochrome-b gene and genotyping of 6 nuclear microsatellite loci were examined to assess genetic diversity and phylogenetic relationship of Ethiopian camels. Examination of 525 individuals revealed significant morphometric differentiation in Afar as compared with the remaining 7 breeds. Analysis of cytochrome-b sequences failed to recover monophyletic groups associated with pastoralist-recognized breeds. Analysis of 6 microsatellite loci from 104 individuals depicted no resolution of distinct genetic lineages in accordance to geographical or designated breeds. Overall, separation of 2 ecotypes based on the morphometric data was supported; however, genetic analysis of cytochrome-b and microsatellite data failed to support any unique genetic lineage or statistically significant population structure.

Variability of Some Milk-Associated Genes and Proteins in Several Breeds of Saudi Arabian Camels.

To gain knowledge on the molecular basis of diversity of several clans of Saudi camel (Camelus dromedarius) characterization of these animals was conducted at both genetic and protein levels. To this end, blood and milk samples were collected from several camel breeds at different Saudi Arabia locations (northern Jeddah, Riyadh, and Alwagh governorates). Genomic DNA was extracted from blood of four Saudi camel breeds (Majahem, Safra, Wadha, and Hamara), and DNA fragments of the casein and α-lactalbumin genes were amplified. The retrieved DNA sequences were analyzed for genetic variability. The inter-simple sequence repeat technique was used for confirming the relationships among the analyzed camel breeds, and the PCR-RFLP with two restriction enzymes was utilized for exploring their molecular variations. The number of haplotypes, gene diversity, nucleotide diversity, average number of nucleotide differences, and sequence conservation were calculated for all the analyzed DNA sequences. These analyses revealed the presence of several single nucleotide polymorphisms in the analyzed DNA sequences. A group of neighbor joining trees was built for inferring the evolutionary variations among the studied animals. Protein profiling of milk from different camel clans was also conducted, and differences between and within the Saudi camel clans were easily found based on the isoelectric focusing (IEF) profiles using ampholytes with different IEF range. This study revealed that analyzed camel breeds show low levels of genetic differences. This may be a reflection of the evolutionary history of C. dromedarius that was domesticated based on a highly homogeneous ancestor ecotype.

Combined hybridization capture and shotgun sequencing for ancient DNA analysis of extinct wild and domestic dromedary camel.

The performance of hybridization capture combined with next-generation sequencing (NGS) has seen limited investigation with samples from hot and arid regions until now. We applied hybridization capture and shotgun sequencing to recover DNA sequences from bone specimens of ancient-domestic dromedary (Camelus dromedarius) and its extinct ancestor, the wild dromedary from Jordan, Syria, Turkey and the Arabian Peninsula, respectively. Our results show that hybridization capture increased the percentage of mitochondrial DNA (mtDNA) recovery by an average 187-fold and in some cases yielded virtually complete mitochondrial (mt) genomes at multifold coverage in a single capture experiment. Furthermore, we tested the effect of hybridization temperature and time by using a touchdown approach on a limited number of samples. We observed no significant difference in the number of unique dromedary mtDNA reads retrieved with the standard capture compared to the touchdown method. In total, we obtained 14 partial mitochondrial genomes from ancient-domestic dromedaries with 17-95% length coverage and 1.27-47.1-fold read depths for the covered regions. Using whole-genome shotgun sequencing, we successfully recovered endogenous dromedary nuclear DNA (nuDNA) from domestic and wild dromedary specimens with 1-1.06-fold read depths for covered regions. Our results highlight that despite recent methodological advances, obtaining ancient DNA (aDNA) from specimens recovered from hot, arid environments is still problematic. Hybridization protocols require specific optimization, and samples at the limit of DNA preservation need multiple replications of DNA extraction and hybridization capture as has been shown previously for Middle Pleistocene specimens.

Genetic diversity and phylogeographic structure of Bactrian camels shown by mitochondrial sequence variations.

The Bactrian camel includes various domestic (Camelus bactrianus) and wild (Camelus ferus) breeds that are important for transportation and for their nutritional value. However, there is a lack of extensive information on their genetic diversity and phylogeographic structure. Here, we studied these parameters by examining an 809-bp mtDNA fragment from 113 individuals, representing 11 domestic breeds, one wild breed and two hybrid individuals. We found 15 different haplotypes, and the phylogenetic analysis suggests that domestic and wild Bactrian camels have two distinct lineages. The analysis of molecular variance placed most of the genetic variance (90.14%, P < 0.01) between wild and domestic camel lineages, suggesting that domestic and wild Bactrian camel do not have the same maternal origin. The analysis of domestic Bactrian camels from different geographical locations found there was no significant genetic divergence in China, Russia and Mongolia. This suggests a strong gene flow due to wide movement of domestic Bactrian camels.

Evolution of camel CYP2E1 and its associated power of binding toxic industrial chemicals and drugs.

Camels are raised in harsh desert environment for hundreds of years ago. By modernization of live and the growing industrial revolution in camels rearing areas, camels are exposed to considerable amount of chemicals, industrial waste, environmental pollutions and drugs. Furthermore, camels have unique gene evolution of some genes to withstand living in harsh environments. In this work, the camel cytochrome P450 2E1 (CYP2E1) is compromised to detect its evolution rate and its power to bind with various chemicals, protoxins, procarcinogens, industrial toxins and drugs. In comparison with human CYP2E1, camel CYP2E1 more efficiently binds to small toxins as aniline, benzene, catechol, amides, butadiene, toluene and acrylamide. Larger compounds were more preferentially bound to the human CYP2E1 in comparison with camel CYP2E1. The binding of inhalant anesthetics was almost similar in both camel and human CYP2E1 coinciding with similar anesthetic effect as well as toxicity profiles. Furthermore, evolutionary analysis indicated the high evolution rate of camel CYP2E1 in comparison with human, farm and companion animals. The evolution rate of camel CYP2E1 was among the highest evolution rate in a subset of 57 different organisms. These results indicate rapid evolution and potent toxin binding power of camel CYP2E1.

The differentiation of camel breeds based on meat measurements using discriminant analysis.

The meat productivity of camel in the tropics is still under investigation for identification of better meat breed or type. Therefore, four one-humped Saudi Arabian (SA) camel breeds, Majaheem, Maghateer, Hamrah, and Safrah were experimented in order to differentiate them from each other based on meat measurements. The measurements were biometrical meat traits measured on six intact males from each breed. The results showed higher values of the Majaheem breed than that obtained for the other breeds except few cases such dressing percentage and rib-eye area. In differentiation analysis, the most discriminating meat variables were myofibrillar protein index, meat color components (L* and a*, b*), and cooking loss. Consequently, the Safrah and the Majaheem breeds presented the largest dissimilarity as evidenced by their multivariate means. The canonical discriminant analysis allowed an additional understanding of the differentiation between breeds. Furthermore, two large clusters, one formed by Hamrah and Maghateer in one group along with Safrah. These classifications may assign each breed into one cluster considering they are better as meat producers. The Majaheem was clustered alone in another cluster that might be a result of being better as milk producers. Nevertheless, the productivity type of the camel breeds of SA needs further morphology and genetic descriptions.

The de novo genome assembly and annotation of a female domestic dromedary of North African origin.

The single-humped dromedary (Camelus dromedarius) is the most numerous and widespread of domestic camel species and is a significant source of meat, milk, wool, transportation and sport for millions of people. Dromedaries are particularly well adapted to hot, desert conditions and harbour a variety of biological and physiological characteristics with evolutionary, economic and medical importance. To understand the genetic basis of these traits, an extensive resource of genomic variation is required. In this study, we assembled at 65× coverage, a 2.06 Gb draft genome of a female dromedary whose ancestry can be traced to an isolated population from the Canary Islands. We annotated 21,167 protein-coding genes and estimated ~33.7% of the genome to be repetitive. A comparison with the recently published draft genome of an Arabian dromedary resulted in 1.91 Gb of aligned sequence with a divergence of 0.095%. An evaluation of our genome with the reference revealed that our assembly contains more error-free bases (91.2%) and fewer scaffolding errors. We identified ~1.4 million single-nucleotide polymorphisms with a mean density of 0.71 × 10(-3) per base. An analysis of demographic history indicated that changes in effective population size corresponded with recent glacial epochs. Our de novo assembly provides a useful resource of genomic variation for future studies of the camel's adaptations to arid environments and economically important traits. Furthermore, these results suggest that draft genome assemblies constructed with only two differently sized sequencing libraries can be comparable to those sequenced using additional library sizes, highlighting that additional resources might be better placed in technologies alternative to short-read sequencing to physically anchor scaffolds to genome maps.

The Arabian camel Camelus dromedarius heat shock protein 90α: cDNA cloning, characterization and expression.

Heat shock protein 90 (Hsp90) is a highly conserved ubiquitous molecular chaperone contributing to assisting folding, maintenance of structural integrity and proper regulation of a subset of cytosolic proteins. In the present study, a heat shock protein 90α full length coding cDNA was isolated and cloned from the Arabian one-humped camel by reverse transcription polymerase chain reaction (RT-PCR). The full length cDNA sequence was submitted to NCBI GeneBank under the accession number KF612338. The sequence analysis of the Arabian camel Hsp90α cDNA showed 2202bp encoding a protein of 733 amino acids with estimated molecular mass of 84.827kDa and theoretical isoelectric point (pI) of 5.31. Blast search analysis revealed that the C. dromedarius Hsp90α shared high similarity with other known Hsp90α. Comparative analyses of camel Hsp90α protein sequence with other mammalian Hsp90s showed high identity (85-94%). Heterologous expression of camel Hsp90α cDNA in E. coli JM109 (DE3) gave a fusion protein band of 86.0kDa after induction with IPTG for 4h.

Analysis of heavy and light chain sequences of conventional camelid antibodies from Camelus dromedarius and Camelus bactrianus species.

Camel antibodies have been widely investigated, but work has focused upon the unique heavy chain antibodies found across camelid species. These are homodimers, devoid of light chains and the first constant heavy chain domain. Camelid species also display conventional hetero-tetrameric antibodies with identical pairs of heavy and light chains; in Camelus dromedarius these constitute 25% of circulating antibodies. Few investigations have been made on this subset of antibodies and complete conventional camel IgG sequences have not been reported. Here we study the sequence diversity of functional variable and constant regions observed in 57 conventional heavy, 18 kappa and 35 lambda light chains of C. dromedarius and Camelus bactrianus. We detail sequences of the full kappa and lambda light chain, variable and CH1 region for IgG1a and IgG1b and the CH2 and CH3 region for IgG1a. The majority (60%) of IgG1 variable region sequences aligned with the human IgHV3 family (clan III) and had leader sequences beginning with MELG whereas the remaining sequences aligned with the IgHV4 (clan II) and had leader sequences beginning with MRLL. Distinct differences in CDR length were observed between the two; where CDR1 was typically 5 and 7 residues and CDR2 at 17 and 16 residues, respectively. CDR3 length of IgHV4 (range 11 to 20) was closer to that typical of VHH antibodies than that of IgHV3 (range 3 to 18 residues). Designed oligonucleotide primers have enabled identification of paired heavy and light chains of conventional camel antibodies from individual B cell clones.

Estimating the population mutation rate from a de novo assembled Bactrian camel genome and cross-species comparison with dromedary ESTs.

The Bactrian camel (Camelus bactrianus) and the dromedary (Camelus dromedarius) are among the last species that have been domesticated around 3000-6000 years ago. During domestication, strong artificial (anthropogenic) selection has shaped the livestock, creating a huge amount of phenotypes and breeds. Hence, domestic animals represent a unique resource to understand the genetic basis of phenotypic variation and adaptation. Similar to its late domestication history, the Bactrian camel is also among the last livestock animals to have its genome sequenced and deciphered. As no genomic data have been available until recently, we generated a de novo assembly by shotgun sequencing of a single male Bactrian camel. We obtained 1.6 Gb genomic sequences, which correspond to more than half of the Bactrian camel's genome. The aim of this study was to identify heterozygous single-nucleotide polymorphisms (SNPs) and to estimate population parameters and nucleotide diversity based on an individual camel. With an average 6.6-fold coverage, we detected over 116 000 heterozygous SNPs and recorded a genome-wide nucleotide diversity similar to that of other domesticated ungulates. More than 20 000 (85%) dromedary expressed sequence tags successfully aligned to our genomic draft. Our results provide a template for future association studies targeting economically relevant traits and to identify changes underlying the process of camel domestication and environmental adaptation.

Cloning and phylogenetic analysis of Interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) from Indian dromedaries (Camelus dromedarius).

The cDNAs of two proinflammatory cytokines viz., IL-6 and TNF-α from dromedarian camels were amplified by PCR using bactrian camel sequences and subsequently cloned for sequence analysis. Relationship based on amino acid revealed that dromedarian camel IL-6 shared 99.5% identity both at nucleotide and amino acid level with bactrian camel IL-6 and in case of TNF-α, the identity of dromedarian camel was 99.4% and 99.1% at nucleotide and amino acid level, respectively with that of bactrian camel. Phylogenetic analysis based on their amino acid sequences indicated the close relationship in these cytokine genes between dromedarian camel and other members of camelids.

Species-specific polymerase chain reaction amplification of camel (Camelus) DNA extracts.

A sensitive polymerase chain reaction (PCR) method based on amplification of a specific DNA fragment was established for the identification of camel (Camelus) materials. The species-specific primer pair L183/H372 was designed based on the nucleotide sequence of the mitochondrial cytochrome b gene, and its specificity was confirmed by amplification of 3 camel (domestic double-humped camel, wild double-humped camel, wild one-humped camel) samples and 11 non-Camelus animal (sheep, goat, pig, chicken, cattle, fish, dog, horse, donkey, deer, and rabbit) materials. An expected 208 base pair fragment was amplified from camel materials; no cross-reactive or additional fragments were generated from other animal materials. Taq I restriction endonuclease digestion of the unpurified PCR product can be used routinely to confirm the camel origin of the amplified sequence.

Genetic diversity and relationships of indigenous Kenyan camel (Camelus dromedarius) populations: implications for their classification.

The genetic diversity and relationships amongst the dromedary (Camelus dromedarius) populations are poorly documented. Four recognized Kenyan dromedary breeds (Somali, Turkana, Rendille, Gabbra) and dromedary from Pakistan and the Arabian Peninsula (Saudi Arabia, United Arab Emirates) were studied using 14 microsatellite loci. Phylogenetic analysis showed that Kenyan dromedaries are distinct from Arabian and Pakistani populations. Expected heterozygosity and allelic diversity values indicate that Kenyan dromedaries are less diverse than non-Kenyan populations. With the exception of the Somali population, the Kenyan dromedaries are poorly differentiated (average FST=0.009), with only one to two loci separating the Gabbra, Rendille and Turkana populations studied (P < 0.05). Individual assignments were performed using the maximum likelihood method. A correct breed assignment of only 39-48% was observed for the Kenyan dromedaries, using an allocation stringency of a log of the odds ratio >2. Our results do not support the present classification of the indigenous Kenyan dromedary into four distinct breeds based on socio-geographical criteria. Instead, our results point to just two separate genetic entities, the Somali and a group including the Gabbra, Rendille and Turkana populations.

[The dromedary: ancestry, history of domestication and medical treatment in early historic times]. / Das Dromedar: Herkunft, Domestikationsgeschichte und Krankheitsbehandlung in frühgeschichtlicher Zeit.

Research on camel bones from archaeological excavations in North Africa and the Near East contradict the current opinion in textbooks that the dromedary and the Bactrian camel are domesticated forms derived from a single species, the two-humped wild camel (C. ferus). The probable ancestor of the domestic one-humped camel is Thomas' camel (C. thomasi). The wild dromedary was probably domesticated on the Arabian peninsula, perhaps as early as the 4th millennium BC. From the 2nd millennium BC onward there is evidence for the use of the one-humped camel for riding and transport purposes. Camel-keeping also implied the therapy of diseased animals, the main source of information about their treatment being medieval arabic texts dealing with veterinary medicine. An expert of such an early 13th century text is presented and discussed. From this it can be seen that the medieval camel treatment fits the principles of the so-called humoral theory, a medical concept which at that time already existed for more than 1500 years.

Molecular evolution of the family Camelidae: a mitochondrial DNA study.

We report the first molecular evolutionary analysis of the family Camelidae by analysing the full DNA sequence of the mitochondrial cytochrome b gene. Estimates for the time of divergence of the Old World (Camelini) and New World (Lamini) tribes obtained from sequence data are in agreement with those derived from the fossil record. The DNA sequence data were also used to test current hypotheses concerning the ancestors of the domesticated llama and alpaca. The results show that hybridization has occurred in the ancestry of both domesticated camelids, obscuring the origin of the domestic species.