Molecular epidemiology of Shigella flexneri isolated from pediatrics in a diarrhea-endemic area of Khyber Pakhtunkhwa, Pakistan.

Shigella flexneri is considered as an important causative agent of Shigellosis causing diarrhea in the countries with a low socioeconomic status. No study has been carried out on the molecular prevalence of S. flexneri in Khyber Pakhtunkhwa, Pakistan. So this study was designed to evaluate the molecular prevalence of S. flexneri and their associated risk factors. A total of 2014 diarrheal stool samples were collected from January 2016 to May 2017 from pediatrics patients of Khyber Pakhtunkhwa followed by identification of S. flexneri through biochemical, serological, and molecular methods. The overall prevalence of Shigella species was found to be 7.9% (n = 160). The predominant Shigella specie was S. flexneri (n = 155, 96.8%) followed by S. boydii (n = 5, 3.1%). Interestingly, no sample was found positive for S. sonnei and S. dysenteriae. The majority of Shigellosis cases occurred from June to September. Potential risk factors related with Shigellosis were unhygienic latrine usage, bad hand washing, and consumption of unhygienic food and water, and pipe leakage in the sewage system. In this study, we have observed a high number of Shigellosis cases especially those caused by S. flexneri. It is suggested that effective health awareness programs should be organized by the regional health authorities to minimize the magnitude of pediatrics Shigellosis.

Natural occurrence of mycotoxins in medicinal plants: a review.

Medicinal plants are widely used as home remedies and raw materials for the pharmaceutical industries. Herbal remedies are used in the prevention, treatment and cure of disorders and diseases since ancient times. However, use of medicinal herbs may not meet the requirements of quality, safety and efficacy. During harvesting, handling, storage and distribution, medicinal plants are subjected to contamination by various fungi, which may be responsible for spoilage and production of mycotoxins. The increasing consumption of medicinal plants has made their use a public health problem due to the lack of effective surveillance of the use, efficacy, toxicity and quality of these natural products. The increase in use of medicinal plants may lead to an increase in the intake of mycotoxins therefore contamination of medicinal plants with mycotoxins can contribute to adverse human health problems and therefore represents a special hazard. Numerous natural occurrences of mycotoxins in medicinal plants and traditional herbal medicines have been reported from various countries including Spain, China, Germany, India, Turkey and from Middle East as well. This review discusses the important mycotoxins and their natural occurrences in medicinal plants and their products.

Incidence of Helminthic and Viral Coinfections in Malaria Patients in the Tertiary Care Hospital Setup.

Introduction: This study determines the incidence of common viral and helminth coinfections with malaria in the tertiary care hospital set up in southern Khyber Pakhtunkhwa, Pakistan. Materials and Methods: The multidimensional research included malaria patients admitted to different hospitals of district Kohat during January and December 2021. Stool samples and blood were assembled from the patients. Giemsa-stained microscopy-positive samples were processed by the immunochromatography technique (ICT) to identify Plasmodium species. Common viral infections such as viral hepatitis (A, B, and C), HIV, and dengue (DENV) were analyzed by ICT kits while SARS-CoV-2 was confirmed through real-time PCR. Furthermore, the intestinal helminths were identified using the Kato-Katz thick smear method. Results: Among 1278 patients, 548 were diagnosed with malaria, 412 (75.2%) were positive for P. vivax infection, 115 (21%) for P. falciparum, and 21 (3.8%) for mixed malaria infection (P. vivax/P. falciparum), with a higher incidence among males (65.2%) than females (34.8%). Coinfection with helminths was positive in 215 (39.3%) malaria patients. The most common infections were caused by the Ascaris lumbricoides species (42.6%) followed by Enterobius vermicularis (31.7%) and hookworm. A total of 24.6% of malaria-positive cases were also coinfected with different viruses with higher frequencies of confection for HAV (8.2%) and DENV (6.2%), respectively. The patients revealed higher incidence of coinfections with P. falciparum (57%) as compared with P. vivax (39.2%) and mixed infections (3.7%). Conclusion: This study demonstrated that the study population exhibited a significant incidence of coinfections with intestinal helminth and viral malaria.

Characterization and Antibacterial Potential of Iron Oxide Nanoparticles in Eradicating Uropathogenic E. coli.

Proper management and control measurements are needed to stop the spread of highly pathogenic E. coli isolates that cause urinary tract infections (UTI) by developing new antibacterial agents to ensure the safety of public health. Therefore, the present investigations were used to achieve the synthesis of iron oxide nanoparticles (IONPs) via a simple coprecipitation method using ferric nitrates Fe (NO3)3 as the precursor and hydrazine solution as the precipitator and to explore the antibacterial activity against eradicating Uropathogenic Escherichia coli (E. coli). The synthesized IONPs were further studied using a UV-vis spectrophotometer, Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), and scanning electron microscopic (SEM) analysis. The maximum surface plasmon resonance peak was observed as absorption at 320 nm in a colloidal solution to validate the synthesis of IONPs. The FT-IR analysis was used to identify different photoactive functional groups that were responsible for the reduction of Fe (NO3)3 to IONPs. The crystalline nature of synthesized IONPs was revealed by XRD patterns with an average particle size ranging as 29 nm. The SEM image was employed to recognize the irregular morphology of synthesized nanoparticles. Moreover, significant antibacterial activity was observed at 1 mg/mL stock solution but after (125, 250, and 500 µg/mL) dilution, the synthesized IONPs showed moderate activity and became inactive at lower concentrations. The morphological and biochemical tests were used to confirm the presence of E. coli in the samples. Furthermore, the minimum inhibitory concentration (MIC) and minimum bacterial concentration (MBC) were carried out to determine the inhibitory concentrations for the isolated bacteria. The isolated E. coli were also subjected to antibiotic sensitivity testing that showed high resistance to antibiotics such as penicillin and amoxicillin. Thus, the findings of this study were to use IONPs against antibiotic resistance that has been developed in an inappropriate way.

Antimicrobial Resistance and Associated Risk Factors for Clostridium difficile in Patients Attending Tertiary Care Settings.

To determine the incidence of antimicrobial-resistant emerging pathogens, Clostridium difficile, and its associated risk factors in tertiary care setups of Pakistan. This cross-sectional prospective study was conducted from January 2019 to December 2020, to determine the prevalence and antimicrobial resistance patterns of C. difficile strains isolated from 450 stool specimens of patients suffering from diarrhea hospitalized in tertiary care hospitals in Peshawar, Pakistan. The stool samples of the patients were processed for culture and detection of toxin A and toxin B by enzyme-linked immunosorbent assay (ELISA) and tpi PCR. The drug sensitivity test was performed for antibiotics including ampicillin, cefixime, cefepime, amoxicillin, nalidixic acid, sulpha/TMP (SXT), chloramphenicol, metronidazole, vancomycin, ciprofloxacin, levofloxacin, and imipenem. Of 450 stool specimens, 108 (24%) were positive for C. difficile by stool culture, whereas 115 (25.5%) were only positive for C. difficile toxins based on ELISA and PCR (128 (28.6%). Of 108, 90.7% (n = 98) isolates were resistant to one antibiotic, and 90 (83.4%) were resistant to three or more antimicrobials. The highest resistance rates were found against penicillin (83.3%) followed by amoxicillin (70%), nalidixic acid (61%), and metronidazole (38%), and the lowest resistance was found against vancomycin (6.4%) and imipenem (3.7%). CDI was statistically significantly correlated with increased age, use of antibiotics, abdominal surgeries, use of proton pump inhibitors and H2a, and presence of comorbidities. The high frequency of C. difficile in Peshawar, Pakistan, indicates that CDI is an important nosocomial infection in different hospitals. The results will be helpful for clinicians to redesign control and therapeutic strategies in hospitals.

Biological Control of Hyalomma Ticks in Cattle by Fungal Isolates.

Ticks pose a major threat to cattle health and production in Pakistan because they transmit pathogens of diseases like Babesiosis and Theileriosis. Hyalomma spp., found across Africa, Asia, and Europe, are especially problematic. This study explored biocontrol of Hyalomma spp. using spore-free fungal culture filtrates collected from dairy farm soil in Kohat, Pakistan. Three fungal species of the genera Alternaria, Aspergillus, and Penicillium were isolated, and their filtrates were tested against tick adults and larvae. Filtrate concentrations were prepared at different strengths. Data were taken after the exposure of adults and larvae ticks to various concentrations of the fungal filtrates. Results indicated that at 100% concentration, all fungal filtrates induced 100% mortality in adults and larvae. Decreasing filtrate concentration lowered tick mortality. The lowest concentration caused the least mortality. The effect was time- and dose-dependent. In conclusion, spore-free fungal culture filtrates can provide biocontrol of Hyalomma spp. in a time- and concentration-dependent manner. Further research should explore the active compounds causing mortality and optimal application methods. The process outlined here provides a natural biocontrol alternative to chemical pesticides to reduce tick infestations and associated cattle diseases in Pakistan.

Whole Exome Sequence Analysis for Inborn Errors of IL-12/IFN-γ Axis in Patient with Recurrent Typhoid Fever.

Background: The IL-12/IFN-γ axis pathways play a vital role in the control of intracellular pathogens such as Salmonella typhi. Objective: The study is aimed at using whole exome sequencing (WES) to screen out genetic defects in IL-12/IFN-γ axis in patients with recurrent typhoid fever. Methods: WES using next-generation sequencing was performed on a single patient diagnosed with recurrent typhoid fever. Following alignment and variant calling, exomes were screened for mutations in 25 genes that are involved in the IL-12/IFN-γ axis pathway. Each variant was assessed by using various bioinformatics mutational analysis tools such as SIFT, Polyphen2, LRT, MutationTaster, and MutationAssessor. Results: Out of 25 possible variations in the IL-12/IFN-γ axis genes, only 2 probable disease-causing mutations were identified. These variations were rare and include mutations in IL23R and ZNFX I. Other pathogenic mutations were found, but they were not considered likely to cause disease based on various mutation predictors. Conclusion: Applying WES to the patient with recurrent typhoid fever detects variants that are not much important as other genes in the IL-12/IFN-γ axis. Results of the current study suggest that a large population sizes would be needed to examine the functional relevance of IL-12/IFN-γ axis genes with recurrent typhoid fever.

Molecular detection of rifampicin resistance by GeneXpert® assay among treated and untreated pulmonary tuberculosis patients from Khyber Pakhtunkhwa, Pakistan.

OBJECTIVES: Drug-resistant tuberculosis (TB) is an emerging health problem. Rifampicin (RIF) is the major first-line drug against TB. RIF resistance can be used as a marker for the detection of multidrug-resistant TB (MDR-TB). The purpose of this study was to determine the RIF resistance pattern of Mycobacterium tuberculosis complex isolates among treated and untreated patients in Khyber Pakhtunkhwa, Pakistan. METHODS: A total of 349 drug-treated and untreated TB-diagnosed patients were enrolled in this study. RIF resistance was detected using a GeneXpert® assay for amplification of the RIF resistance-determining region (RRDR) region of the rpoB gene. RESULTS: The overall prevalence of RIF resistance was 5.2% (18/349). Among 49 untreated TB patients, 3 samples (6.1%) were found resistant to RIF. Among 235 patients with a category 1 treatment regimen, 10 samples (4.3%) were resistant to RIF, whilst among 65 patients with a category 2 (Cat-2) treatment regimen, 5 samples (7.7%) were resistant to RIF. A comparison based on patient sex revealed high RIF resistance among male compared with female patients. RIF resistance was highest (4/21; 19.0%) in the 21-40 years age group among Cat-2 patients. CONCLUSIONS: The overall prevalence of RIF resistance was high among treated and untreated TB patients. These findings will be helpful for better monitoring and management of RIF resistance in TB patients from Khyber Pakhtunkhwa, Pakistan.

First report on molecular characterization of Leishmania species from cutaneous leishmaniasis patients in southern Khyber Pakhtunkhwa province of Pakistan.

OBJECTIVE: To report presence of Leishmania major in Khyber Pakhtunkhwa of Pakistan, where cutaneous leishmaniasis (CL) is endemic and was thought to be caused by Leishmania tropica only. METHODS: Biopsy samples from 432 CL suspected patients were collected from 3 southern districts of Khyber Pakhtunkhwa during years 2011-2016. Microscopy on Giemsa stained slides were done followed by amplification of the ribosomal internal transcribed spacer 1 gene. RESULTS: Leishmania amastigotes were detected by microscopy in 308 of 432 samples (71.3%) while 374 out of 432 samples (86.6%) were positive by ribosomal internal transcribed spacer 1 PCR. Subsequent restriction fragment length polymorphism confirmed L. tropica in 351 and L. major in 6 biopsy samples. CONCLUSIONS: This study is the first molecular characterization of Leishmania species in southern Khyber Pakhtunkhwa. It confirmed the previous assumptions that anthroponotic CL is the major CL form present in Khyber Pakhtunkhwa province. Furthermore, this is the first report of L. major from a classical anthroponotic CL endemic focus identified in rural areas of Kohat district in southern Khyber Pakhtunkhwa.

Evaluation of mycotoxins, mycobiota, and toxigenic fungi in selected medicinal plants of Khyber Pakhtunkhwa, Pakistan.

Medicinal plants are used worldwide to treat a variety of ailments. Due to the provenance of medicinal plants, they are subjected to contamination by moulds, which may be responsible for spoilage and production of mycotoxins. The investigation was designed to throw light on mycological and mycotoxicological status of some medicinal plants from Pakistan and the result showed 30 % and 26.7 % samples were contaminated with aflatoxins and ochratoxin A, respectively. Mould contamination was present in 90 % samples, of which 70 % exceeded the permissible limits. Opium poppy, licorice root, and Indian rennet were most contaminated samples. The predominant moulds found were Aspergillus flavus, Aspergillus niger, Aspergillus parasiticus, and Penicillium spp. and 31 % of the 47 isolates tested were found to be toxigenic. The findings indicate that the contamination in the medicinal plants may contribute to adverse human health problems. This information would prove helpful for regulatory agencies to establish limits for these contaminants in medicinal plants and will explore ways for export of herbal products to countries where more stringent permissible limits of mycotoxins exist. The study is first of its kind in the country reporting natural occurrence of mycotoxins in medicinal plants in Pakistan.