Forensic investigation of carcass decomposition and dipteran fly composition over the summer and winter: a comparative analysis of indoor versus outdoor at a multi-story building.

This study aimed to explore the rate of decomposition of rabbit carcasses and the succession pattern of the associated dipteran flies outdoor, indoor, and on the roof of a 4-story building during the summer and winter. A total of 6,069 flies were recorded, with 30.91% reported as 2 waves outdoor and on the roof in the summer and 69.09% as 4 waves outdoor in the winter. The roof showed the most flies in the summer but the least in the winter, whereas the outdoor showed the most in the winter but the least in the summer. The ground and first floors showed the most indoor flies, while the second and third floors showed the least in both seasons. Indoor carcasses decomposed slower than those outdoor, and those on the second and third floors decomposed slower than those on the ground and first floors. Ten fly species from 8 families were identified in the winter, compared to 6 from 5 families in the summer. The most abundant species was Musca domestica Linnaeus (Muscidae) on the roof in the summer, while it was Chrysomya albiceps (Wiedemannn) (Calliphoridae) outdoor in the winter. The rare species (singletons) were Musca sp. (Muscidae) and Megaselia scalaris (Loew) (Phoridae) on the first floor in both seasons, Scaptomyza pallida (Zetterstedt) (Drosophilidae) on the ground floor in the summer, and Atherigona orientalis Schiner (Muscidae) outdoor in the winter. These data highlight the variance in carcass decomposition and fly composition across outdoor, indoor, and the roof of human dwellings, which could be of forensic importance.

Forensic analysis of mosquito blood meal digestion process and the impact of heroin opiate: determination of the post-feeding interval as a PMI estimation.

Females of some mosquito species are anthropophilic, as they feed on human blood to support egg production and, hence, are forensically valuable if found at a crime scene. The present study investigated the blood meal digestion process in Culex pipiens L. both with and without heroin and proposed a method for estimating the post-feeding interval (PFI). Mosquitoes were fed on a control mouse, a heroin-injected mouse, or in vitro heroin-treated mouse blood. The blood meal digestion was then investigated at different hours post-feeding. Data showed that the blood meal size ingested by control mosquitoes was 0.681 ±â€…0.04 mg/mosquito and was completely digested within 45 h post-feeding. An estimation of the PFI was proposed in terms of the rate of hemoglobin (Hb) digestion. The blood meal size of the mosquitoes fed on the in vitro heroin-treated blood and the heroin-injected mouse was 0.96 ±â€…0.06 and 0.79 ±â€…0.01 mg/mosquito and was completely digested within 50 and 55 h post-feeding, respectively. The digestion of Hb started similarly in all experimental mosquitoes until 10 h post-feeding, after which it significantly decreased in heroin-treated blood meals compared with the control ones. This may suggest that heroin impacted the digestion process, as it took an extra 5-10 h to complete. These findings could be valuable in the forensic context since an estimation of PFI is proposed as a potential estimation of the postmortem interval (PMI). However, care should be taken as heroin in the host blood has significantly impacted the overall digestion process and, hence, may bias the PFI/PMI estimation.

A preliminary investigation of rabbit carcass decomposition and attracted ants (Hymenoptera: Formicidae) on the seaward coastal beach of Al-Jubail City, Saudi Arabia.

The current study was carried out in the seaward coastal beach environment of Al-Jubail City, Saudi Arabia, to analyze the rabbit carcass decomposition process, the succession pattern of associated ants, and their potential utility in forensic investigation. Experiments were conducted over a 4-season course (from autumn 2018 to summer 2019). A total of 9 species belonging to the 2 subfamilies, Myrmicinae and Formicinae, were recorded. The myrmicine species were Crematogaster aegyptiaca Mayr, 1862; Messor ebeninus Santschi, 1927; Messor foreli Santschi, 1923; and Monomorium abeillei Andre, 1881. The formicine species were Camponotus xerxes Forel, 1904; Cataglyphis albicans (Roger, 1859); Cataglyphis hologerseniCollingwood & Agosti, 1996; Cataglyphis viaticoides (André, 1881); and Nylanderia jaegerskioeldi (Mayr, 1904). M. abeillei was the only species recorded in all 4 seasons, while M. abeillei and C. albicans were the dominant species in summer and C. aegyptiaca and C. albicans in spring. Diversity was lowest in the autumn, with only 4 species recorded. The COI gene sequences of 5 species have been successfully deposited in the GenBank database for the first time. In total, 4 carcass decomposition stages were observed, with the longest duration in winter (13 days), the shortest in summer (11 days), and in between for both autumn and spring. Most ant species were present during both decay and dry stages, while M. abeillei, C. aegyptiaca, M. ebeninus, and C. albicans were observed in all decomposition stages. These data may indicate that ants on this coastal beach showed seasonal and geographical succession patterns that could be taken into consideration in forensic investigations.

Comparative molecular profiling of antimicrobial resistance and phylogenetic characterization of multidrug-resistant Escherichia coli isolated from meat sources in 2009 and 2021 in Japan.

The global spread of antimicrobial resistance (AMR) is alarming. Escherichia coli is a Gram-negative bacterium that causes healthcare-associated infections and is a major threat to public health. Currently, no comprehensive antimicrobial surveillance of multidrug-resistant E. coli of diverse phylogroups along the meat value chain has been implemented in Higashihiroshima, Japan. Therefore, by employing the One Health approach, 1183 bacterial isolates, including 303 recovered from meat samples in 2009, were screened for the presence of antimicrobial resistance determinants using multiplex PCR and DNA sequencing techniques. Seventy-seven non-duplicate E. coli isolates that harbored AMR genes were subjected to antimicrobial susceptibility testing and the detection of integrons. Phylogenetic characterization, which has not been previously investigated, was used to assign E. coli to one of the eight phylogroups. Twenty-six out of 33 (78.8%) and 34 out of 44 (77.3%) E. coli isolates from 2009 and 2021 exhibited multidrug resistance (MDR) phenotypes, respectively. The most common clinical resistance was observed against ampicillin, tetracycline, kanamycin, sulfamethoxazole/trimethoprim, cefotaxime, and chloramphenicol. Overall, 22.1% (17/77) of the E. coli isolates carried extended-spectrum ß-lactamase (ESBL)-encoding genes and showed the ESBL-resistant phenotypes. For the two isolation years, AmpC/ESBL prevalence decreased from 42.4% in 2009 to 20.5% in 2021. The identified AMR genes included blaCTX-M-1, blaCTX-M-2, blaCTX-M-14, blaCTX-M-15, and blaSHV-12 (ESBL-types); blaSHV-1, blaTEM-1, blaTEM-135, and blaTEM-176 (narrow-spectrum types); blaCMY-4, blaADC-32, blaADC-216, blaACT-48, and blaACT-51 (AmpC types); and integrons. All E. coli isolates were negative for carbapenemase-encoding genes, whereas one isolate from 2009 carried mcr-5.1 allele. Approximately 52% of E. coli isolates identified in 2009 were assigned to phylogroup A compared to the 20.5% in 2021. Notably, the highest proportions of E. coli phylogroups exhibiting MDR were groups A, B1, and F, suggesting that members of these groups are mostly associated with drug resistance. This study highlights the role of meat as a significant reservoir of MDR E. coli and potential source for transmission of AMR genes. Our findings emphasize the importance of continuous monitoring to track the changes in the spread of antimicrobial resistance in the food chain.

Impact of Envenomation With Snake Venoms on Rabbit Carcass Decomposition and Differential Adult Dipteran Succession Patterns.

The current study investigates the postmortem successional patterns of necrophagous dipteran insects and the rabbit carcass decomposition rate upon envenomation with snake venom. In total, 15 rabbits, Oryctolagus cuniculus domesticus L. (Lagomorpha, Leporidae), were divided into 3 groups (5 rabbits each; n = 5); the first and second groups were injected with lethal doses of venoms from the Egyptian cobra, Naja haje L. (Squamata, Elapidae), and the horned viper, Cerastes cerastes L. (Squamata, viperidae), respectively. The third group (control) was injected with 0.85% physiological saline and euthanized with CO2. The carcass decomposition stages: fresh, bloating, decay, and dry were recorded and monitored. Data revealed that envenomation shortened the decomposition process by 3 d, 20% shorter than the control. The overall succession pattern of fly species revealed a lower abundance during the fresh stage, which peaked during the decay stage, and declined to the minimum number in the dry stage at the end of the 15-d experimental duration. A total of 2,488 individual flies, belonging to 21 species of 10 families, were collected from all experimental carcasses. The Calliphoridae, Muscidae, and Sarcophagidae were the most abundant and diverse families, whereas the other seven families were rare and least abundant. Although C. cerastes venom was significantly less lethal than N. haje, it showed a faster carcass decomposition process and a higher impact on fly abundance. These data showed that envenomation impacts insect succession and carcass decomposition, which should be taken into account when using insects in forensic investigations since envenomation with snake venoms is one of the leading causes of death worldwide.

Evaluation of Insecticidal Potentials of Five Plant Extracts against the Stored Grain Pest, Callosobruchus maculatus (Coleoptera: Bruchidae).

Plant based insecticides are considered among the most economic and ecofriendly chemicals for the protection of plants and stored grains. The cowpea weevil (Callosbruchus maculatus) causes more than 90% damage to sored grains in three to six months. The current study investigates insecticidal potentials of five selected botanicals: Melia azedarach, Nicotiana rustica, Azadirachta indica, Nicotiana tabacum and Thuja orientalis. They are explored at six different concentrations (0.5, 1.0, 1.5, 2.0, 2.5 and 3.0%) against C. maculatus and compared to effects of distilled water which is used as a control. Toxicities of 3%(V/V) extracts of N. tabacum, N. rustica, A. indica and T. orientalis against C. maculatus were 100%, 86.11%, 80.56% and 72.22%, respectively. Maximum mortality was caused by N. tabacum and N. rustica (100%), followed by A. indica (82%), whereas minimum mortality was observed in T. orientalis (64%) at 2.5%. Several phytochemicals, alkaloids, saponins, diterphenes, phytosterol, flavonoids and phenols were identified in N. tabacum and N. rustica, while few were present in A. indica. Phytosterol was present in greatest abundance. Saponins were only detected in aqueous extracts of N. rustica and N. tabacum. Taken together, these results indicate the utility of N. tabacum, N. rustica and A. indica as potential botanicals to control pest beetle and cowpea weevil.

Chemical Composition and Evaluation of Antifungal and Insecticidal Activities of Essential Oils Extracted from Jambosa caryophyllus (Thunb.) Nied: Clove Buds.

Jambosa caryophyllus has been used in traditional phytotherapy as a treatment against infections. In the present work, essential oils extracted from clove buds (Jambosa caryophyllus ) (EO-JC) were investigated for their composition, antifungal, and insecticidal properties. Extraction of EO-JC was performed by use of hydrodistillation using a Clevenger-type apparatus, and the EOs were analyzed by gas chromatography coupled with mass spectrometry (GC-MS). Antifungal activity of EO-JC was evaluated by the use of solid-state diffusion (disc method) and microdilution to determine the minimum inhibitory concentration (MIC), against three strains of fungus, Aspergillus niger, Aspergillus flavus, and Fusarium oxysporum. Insecticidal activity of EO-JC against the cowpea weevil, Callosobruchus maculatus, was determined to assess utility of EO-JC to control this pest. Several exposures including inhalation and contact were used to determine lethality, as well as the repulsion test was conducted at concentrations of 4, 8, 16, and 32 µL EO-JC. Characterization of EO-JC by GC/MS revealed 34 compounds accounting for 99.98% of the mass of the extract. The predominant compound was eugenol (26.80%) followed by ß-caryophyllene (16.03%) and eugenyl acetate (5.83%). The antifungal activity of EO-JC on solid media exhibited inhibitions in the range of 49% to 87%, and MIC was between 3.125 and 7.80 µg EO-JC/mL. Insecticidal activity, as determined by the use of the inhalation test, and expressed as the LD50 and LD95 after 96 hours of exposure was 2.32 and 21.92 µL/L air, respectively. In the contact test, a 96-hour exposure resulted in LD50 and LD95 of 5.51 and 11.05 µL/L of air, respectively. EO-JC exhibited insecticidal activity against fungi and pest chickpea weevil.

Optimal Medical Therapy for Secondary Prevention of Acute Coronary Syndrome: A Retrospective Study from a Tertiary Hospital in Sudan.

Background: Five-medication regimen is recommended for patients after acute coronary syndrome (ACS) as a secondary prevention strategy at discharge to reduce recurrence and improve mortality. Objective: This study aimed to assess prescribing of optimal medical therapy (OMT) as five-medication regimens for secondary prevention at discharge after ACS in Sudan. Methods: A retrospective cohort study was performed at a tertiary hospital located in Wad Medani, Sudan, in the period between January and December 2019. Data were collected from patients' files. OMT was defined as a combination of five medications; aspirin and P2Y12 inhibitors, statins, beta-blockers (BBs), and angiotensin-converting enzyme inhibitors (ACEIs)/angiotensin receptor blockers (ARBs) or if a valid contraindication was documented. Results: Of the 619 patients throughout the study period, 591 were selected based on inclusion and exclusion criteria. The median age of patients was 60 years, and 58.9% of patients were male. Diabetes (44.5%) and hypertension (42%) were the most common risk factors. Most patients (58.4%) were diagnosed with ST-segment elevation myocardial infarction. About 99.7% of patients were on aspirin, 99.5% on statins, 97% on clopidogrel, 96.8% on dual antiplatelet therapy, 70.4% on BBs, and 57.9% on ACEIs/ARBs. OMT for secondary prevention was prescribed to 267 (45.2%) patients with ACS at discharge. Conclusion: Although prescriptions for all five guideline-recommended medications after ACS were suboptimal, the study showed a positive trend in prescribing most individual medications.

High Prevalence of ESBL and Plasmid-Mediated Quinolone Resistance Genes in Salmonella enterica Isolated from Retail Meats and Slaughterhouses in Egypt.

The emergence and spread of multidrug-resistant Salmonella enterica (S. enterica) to humans through food of animal origin are considered a major global public health concern. Currently, little is known about the prevalence of important antimicrobial resistance genes in S. enterica from retail food in Africa. Therefore, the screening and characterization of the extended-spectrum ß-lactamase (ESBL) and plasmid-mediated quinolone resistance (PMQR) genes in S. enterica isolated from retail meats and slaughterhouses in Egypt were done by using PCR and DNA sequencing techniques. Twenty-eight out of thirty-four (82.4%) non-duplicate S. enterica isolates showed multidrug-resistance phenotypes to at least three classes of antimicrobials, and fourteen (41.2%) exhibited an ESBL-resistance phenotype and harbored at least one ESBL-encoding gene. The identified ß-lactamase-encoding genes included blaCTX-M-1, blaCTX-M-3, blaCTX-M-13, blaCTX-M-14, blaCTX-M-15, and blaSHV-12 (ESBL types); blaCMY-2 (AmpC type); and blaTEM-1 and blaOXA-1 (narrow-spectrum types). PMQR genes (included qnrA, qnrB, qnrS, and aac(6')-Ib-cr) were identified in 23 (67.6%) isolates. The presence of ESBL- and PMQR-producing S. enterica with a high prevalence rate in retail meats and slaughterhouses is considered a major threat to public health as these strains with resistance genes could be transmitted to humans through the food chain.

Molecular Analysis of Antimicrobial Resistance among Enterobacteriaceae Isolated from Diarrhoeic Calves in Egypt.

The present study was designed to investigate the presence of genes that conferred resistance to antimicrobials among Enterobacteriaceae that were isolated from diarrhoeic calves. A total of 120 faecal samples were collected from diarrhoeic calves that were raised in Kafr El-Sheikh governorate, Egypt. The samples were screened for Enterobacteriaceae. A total of 149 isolates of bacteria were recovered and identified; Escherichia coli was found to be the most overwhelming species, followed by Citrobacter diversus, Shigella spp., Serratia spp., Providencia spp., Enterobacter spp., Klebsiella pneumoniae, Proteus spp., Klebsiella oxytoca, and Morganella morganii. All isolates were tested for susceptibility to 12 antimicrobials; resistant and intermediately resistant strains were screened by conventional polymerase chain reaction for the presence of antimicrobial resistance genes. Of the 149 isolates, 37 (24.8%) exhibited multidrug resistant phenotypes. The most prevalent multidrug resistant species were E. coli, C. diversus, Serratia spp., K. pneumoniae, Shigella spp., Providencia spp., and K. oxytoca. Class 1 integrons were detected in 28 (18.8%) isolates. All isolates were negative for class 2 integrons. The blaTEM gene was identified in 37 (24.8%) isolates, whereas no isolates carried the blaCTX-M gene. The florfenicol gene (floR) was detected in two bacterial isolates (1.3%). The findings of this study reveal that calves may act as potential reservoirs of multidrug resistant bacteria that can be easily transmitted to humans.

Prevalence of extended-spectrum ß-lactamase (ESBL)-producing Salmonella enterica from retail fishes in Egypt: A major threat to public health.

The increase in multidrug-resistant Salmonella enterica and its spread from food to humans are considered a serious public health concern worldwide. Little is currently known about the prevalence of extended-spectrum ß-lactamase (ESBL)-producing S. enterica in fish in Africa. Therefore, this study aimed to investigate the existence of ESBL-producing S. enterica in retail fish in Egypt. In total, 200 fish samples were collected randomly from various retail fish markets in Egypt. S. enterica were detected in 19 (9.5%; 95% CI: 5.8-14.4) of the fish samples analyzed. Of the 19 non-repetitive S. enterica isolates, 18 were serologically categorized into eight S. enterica serovars and a non-typable serovar. All 19 S. enterica isolates (100%) showed multidrug-resistant phenotypes to at least three classes of antimicrobials, and 11 (57.9%) exhibited an ESBL-resistant phenotype and harbored at least one ESBL-encoding gene. The ESBL-producing S. enterica serovars were as follows: Kentucky (3 isolates; 15.8%), Enteritidis (2 isolates; 10.5%), Typhimurium (2 isolates; 10.5%), and 1 isolate (5.3%) each of Infantis, Virchow, Paratyphi B, and Senftenberg. The identified ß-lactamase-encoding genes included ESBL-encoding genes blaCTX-M-3, blaCTX-M-14, blaCTX-M-15, blaSHV-1, blaSHV-2 and blaSHV-12; the AmpC ß-lactamase-encoding gene blaCMY-2; and the narrow-spectrum ß-lactamase-encoding genes blaTEM-1 and blaOXA-1. All S. enterica isolates were negative for carbapenemase-encoding genes. Molecular analysis of plasmid transferability and replicon typing revealed that most plasmids (with ß-lactamase-encoding genes) were transferrable, and the most common incompatibility groups were IncI1, IncA/C, IncHI1, and IncN. To the best of our knowledge, this is the first report for molecular characterization of ESBL-producing S. enterica in fish in Egypt. The occurrence of ESBL-producing S. enterica in retail fish constitutes a potential public health threat with the possibility of transmission of these strains with resistance genes to humans. Such transmission would exacerbate the resistance to an important class of antibiotics commonly used in hospitals to treat typhoid and non-typhoidal Salmonella infections.

Immune investigation of the honeybee Apis mellifera jemenitica broods: A step toward production of a bee-derived antibiotic against the American foulbrood.

Keeping honeybees healthy is essential, as bees are not only important for honey production but also cross-pollination of agricultural and horticultural crops; therefore, bees have a significant economic impact worldwide. Recently, the lethal disease, the American foulbrood (AFB), caused great losses of honeybee and decline of global apiculture. Recent studies have focused on using natural insect-derived antibiotics to overcome recently emerged AFB-resistance to conventional antibiotics. In support of these studies, here we investigate the possibility of producing bee-derived anti-AFB antibiotics from an indigenous honeybee, Apis mellifera jemenitica. The immune responses of the third instar stage were first induced against the standards Micrococcus luteus and Escherichia coli compared with the indigenous Paenibacillus larvae (ksuPL5). Data indicated a strong immune response against M. luteus, E. coli and P. larvae 24 h post-P. larvae-injection as revealed by the detection of lysozyme-like, cecropin-like and prophenoloxidase (PO) activities in the plasma of P. larvae-injected third instars. Nodulation activity against injected P. larvae as early as 4 h and peaking 48 h post-P. larvae injection were observed. Potentially active anti-P. larvae immune peptide fractions purified by high-performance liquid chromatography (HPLC) showed significant in vivo therapeutic effects on P. larvae-infected first instars. Mass spectrophotometric analysis and Orbitrap measurements of P. larvae-injected plasma indicated the expression of PO (Mr: 80 kDa), beta-1,3-glucan-binding protein (Mr: 52 kDa) and serine protease 44 isoform X1 (Mr: 46 kDa). This suggests that one or all of these immune peptides contribute to significant survivorship of P. larvae-infected broods, and could be a valuable clue in the search for honeybee-derived anti-AFB natural therapeutic agents. Further molecular characterization and description of the functional roles of these predicted antimicrobial peptides from both broods and adult honeybee may enrich the arsenal of insect-derived antibiotics of therapeutic purposes.

High Prevalence of Antimicrobial Resistance in Gram-Negative Bacteria Isolated from Clinical Settings in Egypt: Recalling for Judicious Use of Conventional Antimicrobials in Developing Nations.

This study was designed to investigate, at the molecular level, the antimicrobial resistance mechanisms of different antimicrobial resistance genes, including, extended-spectrum ß-lactamases, AmpC ß-lactamases, class 1 and 2 integrons, and plasmid-mediated quinolone resistance genes of Gram-negative bacteria isolated from clinical settings in Egypt. A total of 126 nonduplicate Gram-negative isolates were recovered from different clinical samples taken from hospitalized patients in Egypt in 2014. Antimicrobial susceptibility testing showed that, 93.6% (118/126) of the isolates had a multidrug-resistant phenotype. Interestingly, we reported a high level of antimicrobial resistance nearly for all tested antibiotics; to our knowledge, this is the first report from Egypt indicating very high level of antibiotic resistance in Egypt. Polymerase chain reaction screening and DNA sequencing revealed that, 75.4% (95/126) of the isolates harbored at least one extended-spectrum ß-lactamase-encoding gene, with blaCTX-M being the most prevalent (65.9%), followed by blaSHV (46.8%). The AmpC ß-lactamase, blaCMY, was detected in 7.1% (9/126) of bacterial isolates, with blaCMY-42 being the most prevalent. Class 1 integrons were detected in 50.8% (64/126) of the isolates, and class 2 integrons were detected in 2.4% (3/126) of the isolates. The plasmid-mediated quinolone resistance gene, qnr, was detected in 58.7% (74/126) of the tested isolates, with qnrS being the most prevalent. Several antimicrobial resistance determinants were identified in Egypt for the first time, such as SHV-27, SHV-28, SHV-33, SHV-63, SHV-71, SHV-82, SHV-142, CMY-42, CMY-6, and the new CMY-72 like. This study highlights the importance of the conscious use of conventional antimicrobials to overcome the multidrug resistance problem.

Occurrence of Salmonella genomic island 1 (SGI1) in two African Proteus mirabilis strains isolated from diseased chicken flocks.

Two P. mirabilis strains, PmSHR21 and PmSHR38, were collected from chicken flocks in Sharkia Governorate, Egypt in 2016. The two strains showed multidrug-resistance (MDR) phenotypes and were detected to harbour I) floR and sul1 genes conferring resistance to florfenicol and chloramphenicol, and sulfonamides, respectively, II) a ~1.9 kbp class 1 integron containing aadA2-lnuF genes conferring resistance to spectinomycin and streptomycin, and lincosamides, respectively. Interestingly, the two strains were detected to contain SGI1 variant, SGI1-W and inserted between the 3' end of the chromosomal trmE gene and the hipB/hipA toxin/antitoxin homologue. Fingerprinting by ERIC-PCR of the two poultry strains identified in this study and the two human SGI1-carrying P. mirabilis strains described recently in our study showed identical ERIC-pattern between SGI1-W-carrying poultry and human strains, suggesting that they might be clonally related. The detection of SGI1 and its variants in P. mirabilis isolated from humans and chicken flocks in Egypt clarify the geographical and biological spreading through an inter-transmission pathway. To the best of our knowledge, this is the first study detecting SGI1-positive P. mirabilis isolated from chicken flocks in Africa.

Larvicidal Activities of Indigenous Bacillus thuringiensis Isolates and Nematode Symbiotic Bacterial Toxins against the Mosquito Vector, Culex pipiens (Diptera: Culicidae).

BACKGROUND: The incidence of mosquito-borne diseases and the resistance of mosquitoes to conventional pesticides have recently caused a panic to the authorities in the endemic countries. This study was conducted to identify native larvicidal biopesticides against Culex pipiens for utilization in the battle against mosquito-borne diseases. METHODS: Larvicidal activities of new indigenous Bacillus thuringiensis isolates and crude toxin complexes (TCs) of two nematode bacterial-symbionts, Photorhabdus luminescens akhurstii (HRM1) and Ph. luminescens akhurstii (HS1) that tested against Cx. pipiens. B. thuringiensis isolates were recovered from different environmental samples in Saudi Arabia, and the entomopathogenic nematodes, Heterorhabditis indica (HRM1) and He. sp (HS1) were isolated from Egypt. Larvicidal activities (LC50 and LC95) of the potentially active B. thuringiensis strains or TCs were then evaluated at 24 and 48h post-treatment. RESULTS: Three B. thuringiensis isolates were almost as active as the reference B. thuringiensis israelensis (Bti-H14), and seven isolates were 1.6-5.4 times more toxic than Bti-H14. On the other hand, the TCs of the bacterial symbionts, HRM1 and HS1, showed promising larvicidal activities. HS1 showed LC50 of 2.54 folds that of HRM1 at 24h post-treatment. Moreover, histopathological examinations of the HS1-treated larvae showed deformations in midgut epithelial cells at 24h post-treatment. CONCLUSION: Synergistic activity and molecular characterization of these potentially active biocontrol agents are currently being investigated. These results may lead to the identification of eco-friend mosquito larvicidal product(s) that could contribute to the battle against mosquito-borne diseases.

First report in Africa of two clinical isolates of Proteus mirabilis carrying Salmonella genomic island (SGI1) variants, SGI1-PmABB and SGI1-W.

Two Proteus mirabilis strains, designated PmTAN59 and PmKAF126, were isolated from two different Egyptian cities in 2014 and 2015, respectively. PmTAN59 was isolated from a sputum swab from a pneumonia patient in Tanta University Teaching Hospital. PmKAF126 was isolated from a patient with a diabetic foot infection in a hospital in the city of Kafr El-Sheikh. The two isolates were identified with bacterial small ribosomal RNA (16S rRNA) gene amplification and sequencing and tested for antimicrobial sensitivity with a Kirby-Bauer disk diffusion assay. The two strains were resistant to amoxicillin/clavulante, ampicillin, cefotaxime, cefoxitin, ceftriaxone, chloramphenicol, ciprofloxacin, colistin, gentamicin, kanamycin, nalidixic acid, spectinomycin, streptomycin, sulfamethoxazole/trimethoprime, and tetracycline, but sensitive to aztreonam, imipenem, and meropenem. Molecular characterization was used to map the entire backbone, including the multiple antibiotic resistance (MDR) region, of Salmonella genomic island 1 (SGI1). Both isolates carried a structure similar to SGI1, with two different MDR regions corresponding to SGI1-PmABB in PmTAN59 and SGI1-W in PmKAF126. SGI1-PmABB carried an integron of ~1.5kb with a two-gene cassette, aacCA5-aadA7, which confers resistance to gentamicin, streptomycin, and spectinomycin, whereas SGI1-W carried an integron of ~1.9kb containing aadA2-lnuF, which confers resistance to spectinomycin, streptomycin, and lincosamides. PmKAF126 carried the entire SGI1 sequence, however PmTAN59 carried a SGI1 structure with a deletion in the region from ORF S005 to ORF S009 and accompanied by insertion of IS1359 (1258bp). Furthermore, PmTAN59 carried class 2 integron of ~2.2kb containing dfrA1-sat2-aadA1. An ERIC-PCR analysis detected no clonal relationship between the two strains. Molecular screening for other antimicrobial resistance genes and a plasmid analysis indicated that PmTAN59 carried an IncFIB plasmid type. This strain also carried blaTEM-1 and the plasmid-mediated quinolone-resistance gene qnrA1. However, PmKAF126 carried no plasmids and no resistance gene other than that contained in the MDR region of SGI1 and floR gene conferring resistance to florfenicol. To the best of our knowledge, this is the first report of an SGI1-positive P. mirabilis strain in Egypt or on the entire African continent.

High Carbapenem Resistance in Clinical Gram-Negative Pathogens Isolated in Egypt.

The emergence and spread of carbapenem-resistant gram-negative bacteria poses a serious threat to human health worldwide. Currently, little is known about the molecular mechanisms underlying carbapenem resistance and their prevalence among gram-negative bacteria in Egypt. In this study, we analyzed carbapenemase production in gram-negative bacteria isolated from hospitalized patients in Egypt in 2014. All isolates were subjected to phenotypic and genotypic susceptibility testing for carbapenem resistance. Our results indicated a high level of carbapenem-resistant gram-negative bacteria in Egypt, with 50.8% of the isolates harboring at least one carbapenem resistance gene. OXA-48-like and NDM-1 were the most prevalent carbapenemases, being detected in 49.2%, and 47.7% of carbapenemase-positive isolates, respectively, whereas Verona integron-encoded metallo-ß-lactamase (VIM) was detected in only 26.2% of carbapenemase-positive isolates. This study reports for the first time carbapenemase-producing Serratia marcescens, Morganella morganii, and blaVIM-1-like-producing Pseudomonas aeruginosa in Egypt. It is also the first demonstration of the coexistence of different carbapenemases, being detected in 21.5% of carbapenemase-positive isolates. Effective antibiotic supervision, regional surveillance, and early detection of carbapenemase producers are imperative to prevent their future spread to epidemic levels.

Isolation and characterization of native Bacillus thuringiensis strains from Saudi Arabia with enhanced larvicidal toxicity against the mosquito vector Anopheles gambiae (s.l.).

BACKGROUND: Worldwide, mosquito vectors are transmitting several etiological agents of important human diseases, including malaria, causing millions of deaths every year. In Saudi Arabia, as elsewhere, vector-control is based mostly on chemical insecticides which may be toxic and cause environmental deprivation. Here, to support the development of bio-pesticide alternatives, a study was conducted to identify native Bacillus thuringiensis (Bt) isolates with improved toxicity against the malaria vector, Anopheles gambiae (s.l.). METHODS: Sixty-eight Bt isolates were obtained from 300 soil and other samples collected from 16 sites across Saudi Arabia. Bt identification was based on morphological characteristics of colonies, shape of parasporal crystals and biochemical profiles. After characterization of their mosquitocidal activity, larvicidal strains were described through 16S ribosomal DNA gene sequencing, cry, cyt and chi genes PCR-amplification profiles, and SDS-PAGE protein analyses. RESULTS: Spherical Bt crystals were predominant amongst the 68 isolates (34%), while irregular, bi-pyramidal and spore-attached crystals were found in 32, 13 and 21% of strains, respectively. LC50 and LC90 bioassays showed that 23/68 isolates were larvicidal, with distinct biochemical activity profiles compared to non-larvicidal Bt strains. Eight larvicidal strains showed larvicidal activity up to 3.4-fold higher (LC50 range: 3.90-7.40 µg/ml) than the reference Bti-H14 strain (LC50 = 13.33 µg/ml). Of these, 6 strains had cry and cyt gene profiles similar to Bti-H14 (cry4Aa, cry4Ba, cry10, cry11, cyt1Aa, cyt1Ab, cyt2Aa). The seventh strain (Bt63) displaying the highest larvicidal activity (LC50 = 3.90 µg/ml) missed the cry4Aa and cyt1Ab genes and had SDS-PAGE protein profiles and spore/crystal sizes distinct from Bti-H14. The eight strain (Bt55) with LC50 of 4.11µg/ml had cry and cyt gene profiles similar to Bti-H14 but gave a chi gene PCR product size of 2027bp. No strains harbouring cry2, cry17 + 27, cry24 + 40, cry25, cry29, cry30, or cyt2Ba were detected. CONCLUSION: This study represents the first report of several Saudi indigenous Bt strains with significantly higher larvicidal efficacy against An. gambiae than the reference Bti-H14 strain. The very high toxicity of the Bt63 strain, combined with distinct cry and cyt genes and SDS-PAGE-protein profiles makes it a promising candidate for future applications in mosquito bio-control.