Predicting crystal form stability under real-world conditions.

The physicochemical properties of molecular crystals, such as solubility, stability, compactability, melting behaviour and bioavailability, depend on their crystal form1. In silico crystal form selection has recently come much closer to realization because of the development of accurate and affordable free-energy calculations2-4. Here we redefine the state of the art, primarily by improving the accuracy of free-energy calculations, constructing a reliable experimental benchmark for solid-solid free-energy differences, quantifying statistical errors for the computed free energies and placing both hydrate crystal structures of different stoichiometries and anhydrate crystal structures on the same energy landscape, with defined error bars, as a function of temperature and relative humidity. The calculated free energies have standard errors of 1-2 kJ mol-1 for industrially relevant compounds, and the method to place crystal structures with different hydrate stoichiometries on the same energy landscape can be extended to other multi-component systems, including solvates. These contributions reduce the gap between the needs of the experimentalist and the capabilities of modern computational tools, transforming crystal structure prediction into a more reliable and actionable procedure that can be used in combination with experimental evidence to direct crystal form selection and establish control5.

A data-driven and topological mapping approach for the a priori prediction of stable molecular crystalline hydrates.

Predictions of the structures of stoichiometric, fractional, or nonstoichiometric hydrates of organic molecular crystals are immensely challenging due to the extensive search space of different water contents, host molecular placements throughout the crystal, and internal molecular conformations. However, the dry frameworks of these hydrates, especially for nonstoichiometric or isostructural dehydrates, can often be predicted from a standard anhydrous crystal structure prediction (CSP) protocol. Inspired by developments in the field of drug binding, we introduce an efficient data-driven and topologically aware approach for predicting organic molecular crystal hydrate structures through a mapping of water positions within the crystal structure. The method does not require a priori specification of water content and can, therefore, predict stoichiometric, fractional, and nonstoichiometric hydrate structures. This approach, which we term a mapping approach for crystal hydrates (MACH), establishes a set of rules for systematic determination of favorable positions for water insertion within predicted or experimental crystal structures based on considerations of the chemical features of local environments and void regions. The proposed approach is tested on hydrates of three pharmaceutically relevant compounds that exhibit diverse crystal packing motifs and void environments characteristic of hydrate structures. Overall, we show that our mapping approach introduces an advance in the efficient performance of hydrate CSP through generation of stable hydrate stoichiometries at low cost and should be considered an integral component for CSP workflows.

Influence of Solvent Selection on the Crystallizability and Polymorphic Selectivity Associated with the Formation of the "Disappeared" Form I Polymorph of Ritonavir.

The comparative crystallizability and polymorphic selectivity of ritonavir, a novel protease inhibitor for the treatment of acquired immune-deficiency syndrome, as a function of solvent selection are examined through an integrated and self-consistent experimental and computational molecular modeling study. Recrystallization at high supersaturation by rapid cooling at 283.15 K is found to produce the metastable "disappeared" polymorphic form I from acetone, ethyl acetate, acetonitrile, and toluene solutions in contrast to ethanol which produces the stable form II. Concomitant crystallization of the other known solid forms is not found under these conditions. Isothermal crystallization studies using turbidometric detection based upon classical nucleation theory reveal that, for an equal induction time, the required driving force needed to initiate solution nucleation decreases with solubility in the order of ethanol, acetone, acetonitrile, ethyl acetate, and toluene consistent with the expected desolvation behavior predicted from the calculated solute solvation free energies. Molecular dynamics simulations of the molecular and intermolecular chemistry reveal the presence of conformational interplay between intramolecular and intermolecular interactions within the solution phase. These encompass the solvent-dependent formation of intramolecular O-H...O hydrogen bonding between the hydroxyl and carbamate groups coupled with differing conformations of the hydroxyl's shielding phenyl groups. These conformational preferences and their relative interaction propensities, as a function of solvent selection, may play a rate-limiting role in the crystallization behavior by not only inhibiting to different degrees the nucleation process but also restricting the assembly of the optimal intermolecular hydrogen bonding network needed for the formation of the stable form II polymorph.

Investigation of indoor thermal comfort and energy demand in different locations along the sub-Himalayan belt - A simulation based study.

The sub-Himalayan region extends over 2500 km, extending over several countries. Though the effects of climate change is widely anticipated in the diverse but fragile ecosystem of the Himalayas, very less research has been conducted on the indoor environment of the buildings in these regions. In this study, a pre-validated model of 3-storey concrete residential building was used to study the indoor performance and thermal comfort in the face of climate change in the 8 (eight) different hill towns (hill stations) located from west to the east. Rise in ambient and indoor conditions were evident as a part of climate change with colder locations being affected the most. The thermal comfort assessment using both the climate chamber based PMV model and adaptive models revealed the decrease in cold related discomfort and increase in hot related discomfort. On an overall, the indoor conditions improved in these cold locations. The indoor and outdoor thermal condition and thermal comfort plummeted significantly with latitude and elevation. The heating demand in the future climate reduced by about 50-70 % in warmer locations, while the cooling demand increased by as much as 1000-2000 % in cold locations, respectively. Additionally, it was seen that the thermal environment and comfort both declined more rapidly with elevation in the locations lying in the western Himalayas as compared to those in the eastern Himalayas.

Pharmaceutical Development Challenges in a Beyond Rule of Five Prodrug: Case Study of ABBV-167, Phosphate Prodrug of Venetoclax.

ABBV-167, a phosphate prodrug of BCL-2 inhibitor venetoclax, was recently progressed into the clinic as an alternative means of reducing pill burden for patients in high-dose indications. The dramatically enhanced aqueous solubility of ABBV-167 allowed for high drug loading within a crystalline tablet and, when administered in phase I clinical study, conferred venetoclax exposure commensurate with the equivalent dose administered as an amorphous solid dispersion. In enabling the progression into the clinic, we performed a comprehensive evaluation of the CMC development aspects of this beyond the rule of five (bRo5) prodrug. Adding a phosphate moiety resulted in excessively complex chemical speciation and solid form landscapes with significant physical-chemical stability liabilities. A combination of experimental and computational methods including microelectron diffraction (MicroED), total scattering, tablet colorimetry, finite element, and molecular dynamics modeling were used to understand CMC developability across drug substance and product manufacture and storage. The prodrug's chemical structural characteristics and loose crystal packing were found to be responsible for the loss of crystallinity during its manufacturing, which in turn led to high solid-state chemical reactivity and poor shelf life stability. The ABBV-167 case exemplifies key CMC development challenges for complex chemical matter such as bRo5 phosphate prodrugs with significant ramifications during drug substance and drug product manufacturing and storage.

Sensitivity of Different ACTH and Cortisol Concentration Values in Corticotropin-Releasing Hormone Based Tests in Cushing's Disease.

PURPOSE: In Cushing's disease (CD) patients, the aim of the present study is to confirm sensitivity of several ACTH and cortisol concentration values in different time points, during corticotropin-releasing hormone (CRH) stimulation test and during CRH stimulation following dexamethasone suppression (DEX-CRH) test. METHODS: We retrospectively analyzed cortisol and ACTH concentration increment during CRH and DEX-CRH tests in 23 patients with confirmed CD. Cortisol and ACTH concentrations were determined immediately before, 15 min and 30 min after CRH stimulation. We evaluated the sensitivity of different cutoff values including those reported in previous studies, in the diagnosis of CD. RESULTS: During DEX-CRH test, 15 min serum cortisol concentration of 1.4 µg/dl (38 nmol/L) had a sensitivity of 90.9%, and serum cortisol concentration ≥1.27 µg/dl (35 nmol/L) had a sensitivity of 100%. For plasma ACTH, sensitivity of 100% was obtained using ACTH ≥3.5pmol/L (16 pg/ml) at 30 min. During CRH test, 35% increase from baseline in ACTH concentration had a sensitivity of 72.7%. Twenty percent increase in cortisol 30 minutes after stimulation yielded a sensitivity of 85.7%. The best sensitivity of ACTH and cortisol increment was obtained 15 min after stimulation, using 19% and 9% increase, respectively (sensitivity of 100% and 92.8%, respectively). CONCLUSION: During CRH and DEX-CRH tests, the study findings agree with the good sensitivity of ACTH and cortisol cutoff values suggested in previous studies; yet, other cutoff values may give a higher diagnostic sensitivity.

The Correlation between Proliferative Immunohistochemical Markers and Papillary Thyroid Carcinoma Aggressiveness.

Background and Objectives: Papillary thyroid carcinoma (PTC) is one of the most common malignancies of the endocrine system. In order to improve the ability to predict tumor behavior, several studies have been conducted to search for surrogate prognostic immunohistochemical tumor markers. Objective: To evaluate the correlation between the intensity of different immunohistochemical marker staining in PTC and the risk for extrathyroidal extension and metastases. Materials and Methods: The study comprised patients who underwent hemi- or total thyroidectomy. Thyroid tissues were immunohistochemically stained for different tumor proliferative markers: Minichromosome maintenance proteins 2 (MCM2), Ki-67 labeling index, E-Cadherin, Neuropilin-1 and Metallothionein. The correlation between the intensity of each marker staining and the final diagnosis (benign neoplasm and PTC) and the correlation between the intensity of each staining and tumor extrathyroidal extension and metastases were evaluated. Results: The study included 66 patients. Staining for Metallothionein, E-Cadherin and MCM2 significantly differed between benign neoplasm (n = 22) and thyroid-confined PTC (n = 21) (p = 0.002, 0.004 and 0.005, respectively), between benign neoplasm and PTC with extrathyroidal extension (n = 11) (p = 0.001, 0.006 and 0.01, respectively), and between benign neoplasm and PTC with metastases (n = 12) (p = 0.01, <0.001 and 0.037, respectively). No staining correlated with extrathyroidal extension. The intensity of E-Cadherin staining was significantly lower in PTC with metastases than thyroid confined PTC and PTC with extrathyroidal extension (p = 0.028 and 0.021, respectively). Conclusions: Immunohistochemical staining for Metallothionein, E-Cadherin and MCM2 significantly distinguished between benign thyroid tumor and PTC. E-Cadherin staining significantly and inversely correlated with the presence of metastases.

Overcoming Bioavailability Challenges of Dasabuvir and Enabling a Triple-Combination Direct-Acting Antiviral HCV Regimen through a Salt of Very Weak Acid for Oral Delivery.

Dasabuvir is a non-nucleoside polymerase inhibitor for the treatment of hepatitis C virus (HCV) infection. It is an extremely weak diacidic drug (pKa = 8.2 and 9.2) and a prolific solvate former. Due to its exceedingly low aqueous solubility (≤0.127 µg/mL at pH 1-6.8, dose number of 1.31 × 104), crystalline dasabuvir free acid exhibited poor oral bioavailability in initial animal pharmacokinetic (PK) assessment. This necessitated the development of enabling formulation for human clinical studies to achieve the required therapeutic in vivo concentration of dasabuvir. While salt formation has been widely used to enhance the solubility and dissolution rate of solids, this approach has rarely been applied to develop oral solid dosage forms for acidic drugs as weak as dasabuvir due to concerns of rapid disproportionation and crystallization of its free acid. In this contribution, we detail our efforts in identifying dasabuvir monosodium monohydrate as a drug substance that is stable, manufacturable, and, most importantly, significantly enhances the dissolution and oral absorption of this poorly soluble drug. The oral delivery of dasabuvir through the salt approach has enabled the commercialization of the triple-combination direct-acting antiviral HCV regimen, Viekira Pak. The methodologies and solutions identified in targeted studies to overcome technical challenges encountered along the way (i.e., incorporation of polymers to inhibit crystallization and disproportionation and species mapping to enable salt manufacturing process, etc.) can be applied to other insoluble compounds.

Breakage Assessment of Lath-Like Crystals in a Novel Laboratory-Scale Agitated Filter Bed Dryer.

Agitated filter bed dryer is often the equipment of choice in the pharmaceutical industry for the isolation of potent active pharmaceutical ingredients (API) from the mother liquor and subsequent drying through intermittent agitation. The use of an impeller to promote homogeneous drying could lead to undesirable size reduction of the crystal product due to shear deformation induced by the impeller blades during agitation, potentially causing off-specification product and further downstream processing issues. An evaluation of the breakage propensity of crystals during the initial development stage is therefore critical. A new versatile scale-down agitated filter bed dryer (AFBD) has been developed for this purpose. Carbamazepine dihydrate crystals that are prone to breakage have been used as model particles. The extent of particle breakage as a function of impeller rotational speed, size of clearance between the impeller and containing walls and base, and solvent content has been evaluated. A transition of breakage behaviour is observed, where carbamazepine dihydrate crystals undergo fragmentation first along the crystallographic plane [00l]. As the crystals become smaller and more equant, the breakage pattern switches to chipping. Unbound solvent content has a strong influence on the breakage, as particles break more readily at high solvent contents. The laboratory-scale instrument developed here provides a tool for comparative assessment of the propensity of particle attrition under agitated filter bed drying conditions.

Antibiotic resistance, biofilm production ability and genetic diversity of carbapenem-resistant Pseudomonas aeruginosa strains isolated from nosocomial infections in southwestern Iran.

BACKGROUND: This study was aimed to evaluate the antibiotic resistance, biofilm formation, and genetic diversity of carbapenem-resistant Pseudomonas aeruginosa (CRPA) strains isolated from four types of nosocomial infections (NIs) including urinary tract infection (UTI), ventilator-associated pneumonia (VAP), surgical site infection (SSI), and bloodstream infection (BSI). METHODS AND RESULTS: In total, 115 isolates of NIs-causing P. aeruginosa were collected from NIs. Antibiotic susceptibility testing (AST) was performed using disk diffusion method and minimum inhibitory concentrations. Biofilm formation was tested on 96-well polystyrene microtiter plates (MTP). CRPA isolates were genotyped using multiple-locus variable number of tandem repeat analysis (MLVA). The most resistance and susceptibility rates were observed to amikacin (70.6%) and colistin (96.1%), respectively. Colistin and meropenem were the most active antimicrobial agents in VAP, SSI, and BSI. While, colistin and cefepime were the most active in UTIs. In total, 52.2% (n = 60/115) of P. aeruginosa isolates were carbapenem resistant, of which 95.0%, 55.0%, and 5.0% were multidrug-resistant, extensively drug-resistant, and pandrug-resistant, respectively. There was a significant association between resistance to carbapenem and resistance to other antibiotics except for piperacillin/tazobactam. The biofilm production of CRPA isolates was 95.0%, of which 23.3% were strong biofilm producers. Based on MLVA, there were 34 different types of CRPA isolates classified into three main clusters and 5 sub clusters. CONCLUSION: The association of CRPA with other antibiotic resistance, the high rates of biofilm production, and the high genetic diversity of the isolates may be a warning of the need for a careful surveillance program.

The correlation between testosterone, inflammation and cytokine status in type-2 diabetes men.

Type 2 diabetes mellitus (T2DM) is believed to cause hypogonadism through increasing pro-inflammatory cytokines. Tumour necrosis factor-α (TNF-α) is a primary cytokine associated with T2DM. The study explored the association between total testosterone (TT) level and cytokines status in 53 adult males, 27 T2DM (T2DM group) and 26 non-T2DM (control group). Blood samples evaluated fasting plasma glucose, HbA1c, insulin, HOMA-IR, FSH, LH, TT, prolactin, estradiol, cortisol, cortisol-binding globulin, C-reactive protein and eight cytokines (Interferon-gamma, IL-10, IL-13, IL-17A, IL-4, IL-23, IL-6, TNF-α). Data are presented as a median with interquartile interval. TT concentration was lower in the T2DM group [10.9 nmol/L (7.1-12.2) vs. 12.3 nmol/L (10.7-14.9) in control, p = 0.008]. CRP and cortisol in T2DM patients were higher than in control (p = 0.031 and 0.041 respectively). TT was negatively correlated with HOMA-IR, body mass index (BMI) and FSH (p = 0.028, 0.019 and 0.006 respectively). Multiple linear regression models showed that lower TT values were predictable by a linear combination of the independent variables: TNF-α, BMI and T2DM (p = 0.047, 0.023 and 0.019 respectively). High CRP and cortisol levels in T2DM patients suggest an inflammatory state. TT levels associated with TNF-α suggest a role of this cytokine in the aetiology of hypogonadism in T2DM patients.

Implications of the Conformationally Flexible, Macrocyclic Structure of the First-Generation, Direct-Acting Anti-Viral Paritaprevir on Its Solid Form Complexity and Chameleonic Behavior.

Direct-acting antiviral regimens have transformed therapeutic management of hepatitis C across all prevalent genotypes. Most of the chemical matter in these regimens comprises molecules well outside the traditional drug development chemical space and presents significant challenges. Herein, the implications of high conformational flexibility and the presence of a 15-membered macrocyclic ring in paritaprevir are studied through a combination of advanced computational and experimental methods with focus on molecular chameleonicity and crystal form complexity. The ability of the molecule to toggle between high and low 3D polar surface area (PSA) conformations is underpinned by intramolecular hydrogen bonding (IMHB) interactions and intramolecular steric effects. Computational studies consequently show a very significant difference of over 75 Å2 in 3D PSA between polar and apolar environments and provide the structural basis for the perplexingly favorable passive permeability of the molecule. Crystal packing and protein binding resulting in strong intermolecular interactions disrupt these intramolecular interactions. Crystalline Form I benefits from strong intermolecular interactions, whereas the weaker intermolecular interactions in Form II are partially compensated by the energetic advantage of an IMHB. Like Form I, no IMHB is observed within the receptor-bound conformation; instead, an intermolecular H-bond contributes to the potency of the molecule. The choice of metastable Form II is derisked through strategies accounting for crystal surface and packing features to manage higher form specific solid-state chemical reactivity and specific processing requirements. Overall, the results show an unambiguous link between structural features and derived properties from crystallization to dissolution, permeation, and docking into the protein pocket.

Multiplex polymerase chain reaction detection of Streptococcus pneumoniae and Haemophilus influenzae and their antibiotic resistance in patients with community-acquired pneumonia from southwest Iran.

BACKGROUND: This study aimed to evaluate the occurrence of Streptococcus pneumoniae and Haemophilus influenzae in sputum of patients with community-acquired pneumonia (CAP) using culture and multiplex polymerase chain reaction (M-PCR) methods and to survey the antibiotic resistance patterns of aforesaid isolates. RESULT: In total, 23.9 % (n = 22/92) of sputum samples showed positive results in the culture method. S. pneumoniae and H. influenzae were isolated from 15 (16.3 %) and 7 (7.6%) samples, respectively. Using M-PCR, 44 (47.8 %) samples were positive for S. pneumoniae and H. influenzae. Of these, S. pneumoniae and H. influenzae were detected in 33 (35.8%) and 11 (11.9%) of the sputum samples, respectively. The sensitivity, specificity, and accuracy rates of PCR in detection of S. pneumoniae in comparison with culture method were 100, 76.6, and 83.6%, respectively. While, the sensitivity, specificity, and accuracy rates of PCR in detection of H. influenzae in comparison with culture method were 100, 95.3, and 95.8%, respectively. Out of 11 isolates of H. influenzae, two strains confirmed as H. influenzae type b (Hib) and 3 isolates were type f. However, 6 isolates were non-typable. The co-trimoxazole and amoxicillin/clavulanate were the less effective antibiotics against S. pneumonia and H. influenzae, respectively. Ceftriaxone with 13.3% resistance rates was the most effective antibiotic against S. pneumoniae, while, clarithromycin, ceftriaxone, and gentamicin with resistance rates of 28.6% for each one were the most effective chemicals against H. influenzae isolates. CONCLUSION: In this study, the prevalence of S. pneumoniae was more than H. influenzae using culture and M-PCR methods. The M-PCR provided better efficiency in detecting the bacterial agents in CAP patients compared to culture method. This method can improve the early detection of pathogens contributed to CAP. The drug resistant S. pneumoniae and H. influenzae indicated the need to develop a codified monitoring program to prevent further spread of these strains.

Molecular, Solid-State and Surface Structures of the Conformational Polymorphic Forms of Ritonavir in Relation to their Physicochemical Properties.

PURPOSE: Application of multi-scale modelling workflows to characterise polymorphism in ritonavir with regard to its stability, bioavailability and processing. METHODS: Molecular conformation, polarizability and stability are examined using quantum mechanics (QM). Intermolecular synthons, hydrogen bonding, crystal morphology and surface chemistry are modelled using empirical force fields. RESULTS: The form I conformation is more stable and polarized with more efficient intermolecular packing, lower void space and higher density, however its shielded hydroxyl is only a hydrogen bond donor. In contrast, the hydroxyl in the more open but less stable and polarized form II conformation is both a donor and acceptor resulting in stronger hydrogen bonding and a more stable crystal structure but one that is less dense. Both forms have strong 1D networks of hydrogen bonds and the differences in packing energies are partially offset in form II by its conformational deformation energy difference with respect to form I. The lattice energies converge at shorter distances for form I, consistent with its preferential crystallization at high supersaturation. Both forms exhibit a needle/lath-like crystal habit with slower growing hydrophobic side and faster growing hydrophilic capping habit faces with aspect ratios increasing from polar-protic, polar-aprotic and non-polar solvents, respectively. Surface energies are higher for form II than form I and increase with solvent polarity. The higher deformation, lattice and surface energies of form II are consistent with its lower solubility and hence bioavailability. CONCLUSION: Inter-relationship between molecular, solid-state and surface structures of the polymorphic forms of ritonavir are quantified in relation to their physical-chemical properties.

Novel Physics-Based Ensemble Modeling Approach That Utilizes 3D Molecular Conformation and Packing to Access Aqueous Thermodynamic Solubility: A Case Study of Orally Available Bromodomain and Extraterminal Domain Inhibitor Lead Optimization Series.

Drug design with patient centricity for ease of administration and pill burden requires robust understanding of the impact of chemical modifications on relevant physicochemical properties early in lead optimization. To this end, we have developed a physics-based ensemble approach to predict aqueous thermodynamic crystalline solubility, with a 2D chemical structure as the input. Predictions for the bromodomain and extraterminal domain (BET) inhibitor series show very close match (0.5 log unit) with measured thermodynamic solubility for cases with low crystal anisotropy and good match (1 log unit) for high anisotropy structures. The importance of thermodynamic solubility is clearly demonstrated by up to a 4 log unit drop in solubility compared to kinetic (amorphous) solubility in some cases and implications thereof, for instance on human dose. We have also demonstrated that incorporating predicted crystal structures in thermodynamic solubility prediction is necessary to differentiate (up to 4 log unit) between solubility of molecules within the series. Finally, our physics-based ensemble approach provides valuable structural insights into the origins of 3-D conformational landscapes, crystal polymorphism, and anisotropy that can be leveraged for both drug design and development.

Field study of pedestrians' comfort temperatures under outdoor and semi-outdoor conditions in Malaysian university campuses.

Difficulties in controlling the effects of outdoor thermal environment on the human body are attracting considerable research attention. This study investigated the outdoor thermal comfort of urban pedestrians by assessing their perceptions of the tropical, micrometeorological, and physical conditions via a questionnaire survey. Evaluation of the outdoor thermal comfort involved pedestrians performing various physical activities (sitting, walking, and standing) in outdoor and semi-outdoor spaces where the data collection of air temperature, globe temperature, relative humidity, wind speed, solar radiation, metabolic activity, and clothing insulation data was done simultaneously. A total of 1011 participants were interviewed, and the micrometeorological data were recorded under outdoor and semi-outdoor conditions at two Malaysian university campuses. The neutral temperatures obtained which were 28.1 °C and 30.8 °C were within the biothermal acceptable ranges of 24-34 °C and 26-33 °C of the PET thermal sensation ranges for the outdoor and semi-outdoor conditions, respectively. Additionally, the participants' thermal sensation and preference votes were highly correlated with the PET and strongly related to air and mean radiant temperatures. The findings demonstrated the influence of individuals' thermal adaptation on the outdoor thermal comfort levels. This knowledge could be useful in the planning and designing of outdoor environments in hot and humid regions to create better thermal environments.

Synthesis of Hybrid Organic-Inorganic Hydrotalcite-Like Materials Intercalated with Duplex Herbicides: The Characterization and Simultaneous Release Properties.

In this study, a controlled-release formulation of duplex herbicides, namely, 2,4,5-trichlorophenoxybutyric acid (TBA) and 3,4-dichlorophenoxy-acetic acid (3,4D), was simultaneously embedded into Zn-Al-layered double hydroxides (LDHs). The resulting nanohybrid Zinc-Aluminium-3,4D-TBA (ZADTX) was composed of a well-ordered crystalline layered structure with increasing basal spacing from 8.9 Å to 20.0 Å in the Powder X-ray Diffraction (PXRD) with 3,4D and TBA anions located in the gallery of LDHs with bilayer arrangement. The release of 3,4D and TBA fit the pseudo-second-order model. This duplex nanohybrid possessed a well-controlled release property (53.4% release from TBA and 27.8% release from 3,4D), which was highly effective, requiring the use of a small quantity and, hence, environmentally safer.

Emergence of multidrug-resistant Shigella species harboring extended-spectrum beta-lactamase genes in pediatric patients with diarrhea from southwest of Iran.

Owing to the scarce evidence about the multidrug-resistant (MDR) beta-lactamase-producing Shigella isolates in Iran, this study aimed to evaluate the occurrence of extended-spectrum beta-lactamases (ESBL) and AmpC ß-lactamases in Shigella species collected in the southwest of Iran. This study was conducted on Shigella species isolated from stool samples of pediatric patients aged less than 15 years suffering from diarrhea. These isolates were identified by bacteriology tests, serotyping, and polymerase chain reaction (PCR). The antibiotic resistance was determined by disc diffusion. The production of ESBLs and AmpC was investigated by phenotypic confirmatory tests and PCR. In total, 79 Shigella isolates, including 46.8% (n = 37) of S. flexneri and 53.2% (n = 42) of S. sonnei, were isolated, respectively. The most effective antibiotic was imipenem with 93.7% of susceptibility followed by ampicillin (29.1%), and cotrimoxazole (30.4%).The resistance rates of ceftriaxone, ceftazidime, and cefotaxime were 41.8%, 34.2%, and 41.8%, respectively. Also, a total of 57 (72.2%) isolates showed MDR profiles. The phenotypic tests showed that 43.0% (34/79) of isolates can produce ESBLs, and no one was positive for ApmC. The frequency of blaTEM and blaCTX-M were 30.4% and 32.9%, respectively, while the blaPER, blaSHV, and AmpC genes were not detected. The ESBL-producing isolates had a significant (p-value Ë‚ 0.05) resistance rate against ceftriaxone, ceftazidime, cefotaxime, cefepime, erythromycin, and amikacin. The significant prevalence of MDR Shigella isolates harboring ESBL genes highlights the need for effective surveillance measures to prevent the more spread of drug resistance among species.

Detection of Streptococcus gallolyticus in colorectal cancer and inflammatory bowel disease patients compared to control group in southwest of Iran.

There are several pieces of evidence regarding the role of bacteria, such as Streptococcus bovis/gallolyticus in the etiology of gastrointestinal diseases such as colorectal cancer (CRC) and inflammatory bowel disease (IBD). Therefore, the aim of this study was to detect S. gallolyticus subsp. gallolyticus (Sgg) in fecal samples of CRC and IBD patients by culture and molecular methods, in Ahvaz, southwest of Iran. A total of 106 fecal samples were collected from 22 CRC patients, 44 IBD patients, and 40 healthy individuals. The prevalence of Sgg was investigated by culture and polymerase chain reaction (PCR) with specific primers for sodA gene. The results of the stool culture showed that the overall prevalence of Sgg was 9 (13.6%) out of 66 patients. Meanwhile, the number of Sgg isolated from IBD and CRC patients was 7 (15.9%) and 2 (9%), respectively. The bacteria were not isolated from any of the control groups. On the basis of PCR, S. gallolyticus was detected in 24 (36.4%) out of 66 patients. Meanwhile, the number of IBD patients with positive sodA gene was 15 (34.1%) out of 44 cases. In CRC patients, the sodA gene was detected in 9 (40.9%) of 22 cases. Two (5%) of the specimens in the control group had the sodA gene. According to our results, S. gallolyticus subsp. gallolyticus might be involved in CRC and IBD pathogenesis. More investigation with different samples in the various areas might be shaded light on these results.

Prevalence of enterotoxin-encoding genes among diverse Shigella strains isolated from patients with diarrhea, southwest Iran.

Shigella spp. are a major cause of bacillary dysentery, particularly among children in developing countries such as Iran. This study aimed to investigate the presence of two important Shigella enterotoxins (ShET-1 and ShET-2), encoded by the set and sen genes, respectively, by polymerase chain reaction (PCR) assay among Shigella species isolated from children affected by shigellosis in Ahvaz, southwest of Iran. In this cross-sectional study, from June 2016 to April 2017, altogether 117 Shigella isolates were collected from fecal specimens of children aged <15 years with diarrhea in Ahvaz, southwest Iran. All isolates were identified by standard microbiological and molecular methods. The presence of enterotoxin genes was determined by PCR. The most prevalent isolate was Shigella flexneri (47.9%), followed by Shigella sonnei (41%) and Shigella boydii (11.1%), respectively. Shigella dysenteriae was not detected in patients' samples. The frequencies of set1A, set1B, and sen genes were 5.1% (6/117), 15.4% (18/117), and 76.9% (90/117), respectively. This study provides initial background on the prevalence and distribution of the Shigella enterotoxin genes in Shigella isolates in southwest of Iran. In addition, this study revealed a high prevalence of sen enterotoxin gene in Shigella species.