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Klebsiella pneumoniae is ubiquitous and known to be a notorious pathogen of humans, animals, and plant-based foods. K. pneumoniae is a recognized trafficker of antibiotic resistance genes (ARGs) between and from different ecological niches. A total of 775 samples (n = 775) were collected from September 2017 to August 2019 from humans, animals, and environmental sources by applying the random convenient sampling technique. A total of 120 (15.7%) samples were confirmed as K. pneumoniae. The distribution of K. pneumoniae among humans, the environment, and animals was 17.1, 12.38, and 10%, respectively. Isolates have shown significant resistance against all the subjected antibiotics agents except colistin. ARGs profiling revealed that the highest percentage prevalence (67.5%) of blaCTXffM was estimated in the isolates, and various carbapenem resistance genes that were found in the study were blaNDMff1 (43.3%), blaOXAff48 (38%), and (1.67%) blaKPCff2. Overall, 21 distinct sequence types (ST) and 13 clonal complexes (CCs) were found through the multi-locus sequence typing (MLST) analysis. Taking together, the distribution of multi-drug resistance (MDR) K. pneumoniae clones in the community and associated environment is alarming for the health care system of the country. Health policymakers should consider the role of all the integral parts of humans, animals, and the associated environment intently to cope with this serious public and animal health concern. [ABSTRACT FROM AUTHOR]

Background: Neonatal sepsis is a leading cause of morbidity and early-life mortality worldwide, and previous data have reported the highest neonatal mortality rate in Pakistan.Purpose: The present study aimed to decipher the prevalence of group B Streptococcus (GBS)-associated sepsis, coinfections, and antibiotic susceptibility of isolated microbes in neonates.Methods: Blood samples of 100 cases of neonatal sepsis were subcultured on blood agar, GBS agar, and MacConkey agar for isolation of GBS and suspected microbes.Results: Of 100 neonatal blood samples, 85 blood samples were culture-positive, including mixed culture growth (n = 18), 25 samples as early-onset neonatal sepsis (EONS) and 60 as late-onset neonatal sepsis (LONS). Staphylococcus aureus showed high percent positivity (31.4%), followed by Candida sp (16.5%), GBS (14.1%), Klebsiella (14.1%), Staphylococcus epidermidis (11.8%), Pseudomonas (9.4%), Acinetobacter (9.4%), Esherichia coli (8.2%), and Enterococcus (5.9%). GBS was isolated more frequently from EONS than from LONS with 50% coinfections. Mode of delivery, gender, and respiratory distress in neonates were significantly associated with culture-positive sepsis. GBS isolates were highly (91.7%) susceptible to vancomycin, cefotaxime, and chloramphenicol, followed by penicillin (83.3%), ampicillin, and tetracycline (75%). GBS isolates were resistant to erythromycin, clindamycin, ciprofloxacin, and linezolid.Implications For Practice: Our findings evidenced GBS-associated risk factors and antibiotic susceptibility pattern of neonatal sepsis, which will help clinicians in management, control, and treatment of neonatal sepsis.Implications For Research: The epidemiological evidence of GBS-associated neonatal sepsis, demographic characteristics, risk factor data analysis, and drug resistance pattern has disease prevention implications in neonates in low-income countries including Pakistan. [ABSTRACT FROM AUTHOR]

aima Muzammil,2 Nafeesa Yasmeen,4 Muhammad Aamir Aslam,3 Mohsin Khurshid,2 Muhammad Hidayat Rasool,2 Zulqarnain Baloch11Biomedical Research Center, Northwest Minzu University, Lanzhou 730030, People's Republic of China; 2Department of Microbiology, Government College University Faisalabad, Faisalabad, Pakistan; 3Institute of Microbiology, University of Agriculture Faisalabad, Faisalabad, Pakistan; 4College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, People's Republic of ChinaCorrespondence: Bilal Aslam; Zulqarnain Baloch Email drbilalaslam@gcuf.edu.pk; znbalooch@yahoo.com Introduction: Intense livestock farming practices enforcing the farmers to use antibiotics as food supplements on a routine basis. Aberrant use of antibiotics is associated with the emergence of antibiotics resistance and resistant superbugs. Keeping in view the current scenario, the present study was designed for the first time from Pakistan with a specific aim to estimate the prevalence of the carbapenem-resistant Klebsiella pneumoniae in veterinary settings and the waste in Pakistan. Methods: A total of 138 samples from various veterinary sources were collected by employing a nonprobability sampling technique. Isolation and phenotypic identification of carbapenem-resistant K. pneumoniae were performed according to the CLSI standard. Molecular detection of various antibiotic resistance genes (ARGs) was done through PCR by using specific primers against each ARG. According to the pasture scheme, the multilocus sequence typing (MLST) was performed to characterize the K. pneumoniae sequence types (STs). Results: According to the results of the study, overall 9.4% (13/138) isolates were confirmed carbapenem-resistant K. pneumoniae. Among various carbapenem ARGs particularly, the blaNDM-1 was found in 92.3% (12/13) isolates followed by blaOXA-48 84.6% (11/13). MLST results revealed that overall 3 STs were found in the study which includes ST29, ST11, and ST258. Taking together, this is the first study to our best knowledge which demonstrated the prevalence of carbapenem-resistant K. pneumoniae and its various STs prevalent in veterinary settings and the waste of Pakistan. Conclusion: Based on the above-mentioned facts, we suggested that veterinary settings and waste are the potential source and reservoir of carbapenem-resistant K. pneumoniae, which may be disseminated to the environment and ultimately can affect the public and companion livestock health. [ABSTRACT FROM AUTHOR]

Background: The BK Polyomavirus (BKPyV) and JC polyomavirus (JCPyV) infections are widespread in human population and have been associated with severe kidney and brain disorders, respectively. The viruses remain latent primarily in reno-urinary tract, reactivating only in case of a compromised immune system. The seroepidemiology and molecular prevalence of BKPyV and JCPyV have been widely studied both in healthy and immunocompromised patients worldwide. However, data regarding the prevalence of these viruses in the immunocompetent or apparently healthy Pakistani population is lacking. Herein, we present the first ever report on quantitative prevalence of BKPyV and JCPyV in the peripheral blood of a randomly selected cohort of healthy Pakistani population. Methods: A total of 266 whole blood samples were examined. The subjects were divided into three age groups: ≤ 25 years (young), 26-50 years (middle) and ≥ 51 years (elder). Absolute real time PCR assay was designed to quantify the BKPyV and JCPyV viral copy numbers in the range of 106 to 100 copies/mL. Results: Overall, BKPyV was detected in 27.1% (72/266) individuals while JCPyV in 11.6% (31/266) indicating significant difference (p < 0.005) in the distribution of these two viruses. The prevalence of BKPyV significantly decreased from 51% (49/96) in young age group to 8.2% (7/85) in eldest age group. Whereas, JCPyV positivity rate slightly increased from 8.3% (8/96) in young age group to 11.8% (10/85) in elder age group. The median viral load was calculated as 6.2 log and 3.38 log copies/mL of blood for BKPyV and JCPyV, respectively. Notably, no significant difference in viral load of either of the subtypes was found between different age groups. Conclusion: The current study provides an important baseline data on the prevalence and viral load of circulating BKPyV and JCPyV in Pakistani population. The prevalence and viral load of BKPyV was comparatively higher than JCPyV. The prevalence of BKPyV significantly decreased with increase in age while JCPyV positivity rate slightly increased with increasing age. Viral load of both BKPyV and JCPyV was not correlated with the individual ages. [ABSTRACT FROM AUTHOR]

A total of 1500 serum samples were collected from cattle, buffaloes, and humans. The Rose Bengal plate test (RBPT) and enzyme-linked immunosorbent assay (ELISA) detected Brucella antibodies in 56 (10.18%) and 44 (8%) of the cattle, respectively. In samples collected from buffaloes 9.38% and 6.92% were positive, whereas 14% and 11% of the samples from humans were found to be positive by RBPT and ELISA, respectively. Among village workers a higher prevalence of brucellosis was recorded in females than in males. Abattoir-associated personnel also had a higher incidence of brucellosis. Results of the present study revealed that RBPT and ELISA can be used efficiently for mass screening of Brucella antibodies in both humans and animals, and that people that worked with animals had a higher incidence of brucellosis, indicating that they might have been infected by contact with infected animals and might act as carriers. [ABSTRACT FROM AUTHOR]

Background and Objective: Extrapulmonary tuberculosis occurring in breast is a rare disorder in West but is still present in developing nations where tuberculosis is endemic. Tuberculous Mastitis (TM) is difficult to diagnose as it has vague clinical symptoms and insufficient radiological findings. The objective of this study was to determine the frequency of tuberculous mastitis by accessible modalities for early diagnosis. Methods: After the Institutional Ethical Board approvals, Pathology laboratory archives and medical records of 5000 patients from Ibn-e-Sina Hospital Research Institute, Multan and Multi Test Laboratory, were retrieved from the year 2008 to 2018. A retrospective analysis of fine needle aspiration cytology (FNAC) results of patients who presented with breast lumps was carried out. The interpretation of cytological findings and Ziehl Neelsen stained smears was carried out by cytopathologists and microbiologists respectively. Mycobacterium DNA was detected using polymerase chain reaction (PCR) in selected cases. Results: Out of 5000 patients, tuberculosis was diagnosed in 264 (5.28%) cases on FNAC. Acid fast bacilli (AFB) positive smears were found in 79.5%patients while 20.45% cases were AFB negative. Among AFB negative smears, 38 cases were further analyzed by PCR, among which 32 (84.2%) cases showed PCR positive results for TM. An increased frequency of TM among all inflammatory breast lesions from 18% in 2008 to 47.19% in 2018 was observed. Conclusion: Tuberculous Mastitis is an uncommon form of extra-pulmonary tuberculosis. It has non-specific clinical, radiological and varied histological findings that often mimic breast abscess or carcinoma. FNAC is essential to guide the patient to further tests including histochemistry and PCR that can be used as more sensitive and time effective detection modalities. [ABSTRACT FROM AUTHOR]

Liver diseases are among the major health problems in Pakistan. The present study investigated the mechanism of hepatoprotection by cinnamon, cinnamaldehyde and kaempferol in Acetaminophen (APAP)-induced liver injury. Qualitative phytochemical analysis was performed for standardization of cinnamon ethanolic extract. For in-vivo evaluation, Balb/c mice were administered with cinnamon extract (200 mg/kg i.g.), cinnamaldehyde (10 mg/kg i.g.) and kaempferol (10 mg/kg i.g.) for 14 days followed by administration of APAP (200 mg/kg i.p.). At the end of trial, mice were dissected, and blood, liver and spleen samples were collected for biochemical, histopathological and apoptotic genes expression analysis. Statistical analysis was performed for significance of results. The results showed that the hepatic damage due to APAP administration for 8 hours in mice was apparent with increased severity. Cinnamon extract, cinnamaldehyde and kaempferol pretreatment suggested ameliorative effects on organ injury induced by APAP by decreasing the elevated serum levels of total proteins and bilirubin. In addition, APAP exerted severe alterations on liver histology without affecting spleen histology alongwith upregulation of Bad, Bax and Caspase-3 and downregulation of Bcl-2. However, cinnamon, cinnamaldehyde and kaempferol pretreatment ameliorated APAPinduced liver alterations and apoptosis, possibly through their antioxidant activity. In addition, cinnamaldehyde and kaempferol possessed comparable protective potential at 20-times less dose as compared to cinnamon extract alone, suggesting therapeutic potential at lower dose in APAP-induced liver injury and apoptosis. [ABSTRACT FROM AUTHOR]

Klebsiella pneumoniae is an important pathogen that causes pneumonia and bloodstream infections, especially in neonates and intensive care patients. The carbapenems remain an important therapeutic option for clinicians, particularly against cephalosporin-resistant Gram-negative pathogens. This increased use of carbapenems at clinics has resulted in the evolution and spread of carbapenem-resistant K. pneumoniae. In this study, we isolated six bla K. pneumoniae carbapenemase (KPC)-producing strains belonging to sequence type 258 (ST258) from clinical, environmental, and veterinary sources. Antibiotic susceptibility was performed on these isolates and the genes responsible for extended-spectrum beta-lactamases and metallo-beta-lactamase production were screened. The molecular typing was done using multilocus sequence typing. Isolated strains were resistant to various antibiotic classes, including carbapenems, and carried the carbapenem-resistant gene, blaKPC. All strains were susceptible to tigecycline and colistin. This is the first report detecting K. pneumoniae ST258 strains in Pakistan. [ABSTRACT FROM AUTHOR]

Klebsiella pneumoniae is an important pathogen that causes pneumonia and bloodstream infections, especially in neonates and intensive care patients. The carbapenems remain an important therapeutic option for clinicians, particularly against cephalosporin-resistant Gram-negative pathogens. This increased use of carbapenems at clinics has resulted in the evolution and spread of carbapenem-resistant K. pneumoniae . In this study, we isolated six bla K. pneumoniae carbapenemase (KPC)-producing strains belonging to sequence type 258 (ST258) from clinical, environmental, and veterinary sources. Antibiotic susceptibility was performed on these isolates and the genes responsible for extended-spectrum beta-lactamases and metallo-beta-lactamase production were screened. The molecular typing was done using multilocus sequence typing. Isolated strains were resistant to various antibiotic classes, including carbapenems, and carried the carbapenem-resistant gene, blaKPC. All strains were susceptible to tigecycline and colistin. This is the first report detecting K. pneumoniae ST258 strains in Pakistan.

The present study aimed to decipher the bacterial infections in diabetic foot human patients in Pakistan and the anti-microbial susceptibility for clinical relevance. A total of 30 samples were collected from hospitalized type 2 diabetic patients (men and women) having foot ulcers. The collected samples were cultured on mannitol salt agar, Blood agar and MacConkey's agar and cetrimide agar. Gram staining and specific biochemical tests were performed to identify the invading bacteria. Antibiotic sensitivity and resistance pattern was performed for isolated bacteria by Kirby-Bauer disc diffusion method. In diabetic foot ulcers, most prevalent bacteria were S. aureus with percent positivity of 83% followed by E. coli (66%), K. pneumoniae (40%) and P. aeruginosa (16%). The infected ulcer with poly-microorganisms was 83.4% and the infected ulcer with single isolates was 16.6%. Imipenem was found to be most sensitive antibiotic against Gram positive as well as Gram negative bacterial isolates from diabetic foot ulcer human patients. Gram negative isolates from diabetic foot showed resistance to ampicillin, sulfamethoxazole/trimethoprim, cefotaxime/clavulanate, metronidazole. The diabetic foot ulcers of human patients revealed high prevalence of S. aureus followed by E. coli, K. pneumoniae and P. aeruginosa respectively. Imipenem was found to be the most sensitive antibiotic for all the bacterial isolates from foot ulcers of type 2 diabetic patients. This study suggests imipenem as effective antibiotic for treatment of diabetic foot ulcers against bacteria.

Presence of multiple drug resistant pathogens in hospital waste is a serious public health concern, because it may ultimately be disseminated to the human. Current study was designed with the objective to estimate the occurrence of ESBL-producing K. pneumoniae in hospital settings and waste. For this purpose, cross sectional study for a period of one year was designed and non-probability sampling techniques was used to collect total n=112 samples from various sample sources of hospital waste including ward waste, operation theatre waste, wastewater and hospital sludge. Isolation of the K. pneumoniae was done by using selective agar, biochemical identification of the isolates was done through API 20E kit (bioMérieux, France). Molecular identification of the isolates was done by amplifying 16SrDNA with PCR. According to CLSI guidelines disc diffusion assay was performed for antibiotic susceptibility profiling. PCR of MDR isolates was done for the molecular detection of various ESBL genes. Results of the study showed 17 (15%) percentage prevalence of MDR K. pneumoniae from all 112 collected samples. Among various sample sources wastewater showed the highest percentage (23%) prevalence of MDR K. pneumoniae. In 17 confirmed isolates bla CTX-M and blaCTX-M1 were found in 13 (76%) and 12 (71%) respectively which showed the highest prevalence as compared to all other investigated genes. While bla TEM , bla SHV , bla NDM-1 and bla OXA-48 were found with percentage prevalence 9 (53 %), 1 (6%), 9 (53%) and 6 (35 %) respectively. Whereas bla KPC , bla IMP , bla VIM and bla GIM were not detected in any of the isolate. Taking together, strict rules and regulations should be adopted at public as well as hospital level to restrict the dissemination of antibiotic resistance from hospital environment to humans.