Identification of Missing Probes in rpoB gene in Rifampicin Resistant PTB and EPTB Cases Using Xpert MTB/RIF Assay in Sirmaur, Himachal Pradesh.

INTRODUCTION: India has the highest number of TB (27%) and MDR/RR-TB (24%) cases among the notified TB patients. Xpert MTB/ RIF assay is a fully automated cartridge-based real-time PCR to detect MTB and resistance to rifampicin within two hours using three specific primers and five unique molecular probes to target the rpoB gene. This study was done to detect RR-TB cases and frequency of missing probes, which target mutations in rpoB gene, in the different groups of study population in Sirmaur district of Himachal Pradesh. METHODS: All, pulmonary and extrapulmonary specimens, were processed for AFB microscopy and Xpert MTB/RIF assay to diagnose TB and RR-TB. RESULTS: Xpert detected MTBC in 721 patients. Using AFB microscopy, only 284 samples were positive. Of these MTB positive patients, 671 had pulmonary TB and 50 were EPTB cases. Resistance to RIF was detected in 31 (4.29%) cases of which resistance in presumptive tuberculosis group and presumptive drug resistant tuberculosis was 1.51% and 9.30% respectively. Twentyeight (4.17%) PTB cases and three (6%) EP-TB cases were resistant to RIF. The frequency of probe E was highest (77.41%) and mutation combination of probes C and D and E and D was 3.22%. CONCLUSION: Drug resistance in the MTBC is mainly conferred through point mutations in specific gene targets in the bacterial genome. Molecular assays like Genexpert gives rapid diagnosis and Rifampicin resistance. This study helps to provide baseline data of mutations with in the 81 bp of rpoB gene and stresses the need to further evaluate the mutation patterns in this part of the country.

Rubella outbreak in the union territory of Chandigarh, North India.

Rubella virus outbreaks usually occur when a large numbers of susceptible individuals accumulate. The disease presents clinically with fever and maculopapular rash. The present study reports the investigation of rubella outbreak in a modern and well-planned village near Chandigarh, North India. The blood samples were collected from 39 cases with febrile rash and from 15 age and sex matched healthy controls residing in the same locality and subjected for the detection of Rubella IgM and IgG antibodies by Enzyme linked immunosorbent assay. The throat swabs, urine and blood samples from acute cases were also collected and subjected to RT-PCR using the primers targeting the E1 region. The genetic characterization of the rubella virus was carried out to identify the circulating genotypes. In the present outbreak, 13 laboratory confirmed cases were reported. Rubella IgM antibodies were detected in 12/39 (30.7%) patients. Rubella RNA could be detected in 83.3% (5/6) of urine, 22.2% (2/9) of throat swabs, and 8.3% (1/12) of blood samples. The rubella genotype responsible for the present outbreak was identified as genotype 1a. This outbreak highlights the need for the introduction of rubella vaccine in the National Immunization Programme of India to prevent outbreaks and to aim towards the eradication of this disease. This study reports the presence of genotype 1a in North India for the first time and stresses the need for further molecular work to identify the circulating strains of the virus.

A study on characterization of new bacteriocin produced from a novel strain of Lactobacillus spicheri G2 isolated from Gundruk- a fermented vegetable product of North East India: A novel bacteriocin production from Lactobacillus spicheri G2.

Bacteriocin producing Lactobacillus spicheri G2, isolated from Gundruk - a traditional fermented vegetable product of North East India. L. spicheri G2 identified by morphological, biochemical techniques followed by 16S rRNA gene technique. The 16Sr RNA sequence of bacteriocin producer is registered in NCBI under accession no. JX481912. The bacteriocin producing potential of L. spicheri is being reported for the first time in the present investigation. Bacteriocin of L. spicheri G2 showed strong antagonism against food spoiling and pathogenic bacteria viz. Listeria monocytogenes, Staphlococcus aureus, Clostridium perfringens, Streptococcus mutans, Lactobacillus plantarum, Leuconostoc mesenteroides and Bacillus cereus. Bacteriocin production of L. spicheri G2 was enhanced by optimization of production time, pH of medium and incubation temperature by following one variable at a time method. Maximum bacteriocin activity (2000 AU/ml) was recorded in MRS broth at 34 h with an initial pH of 4.0 after incubating at 35 °C. The bacteriocin was purified by single step gel exclusion chromatography. Molecular weight of this novel bacteriocin was determined by SDS PAGE which was found to be 43 kDa. Purified bacteriocin was found resistant to high temperature and varied pH range but sensitive to proteolytic enzymes like trypsin and proteinase k, the characters desirable for food preservation.

Purification and Characterization of Bacteriocin Produced by Lactobacillus brevis UN Isolated from Dhulliachar: a Traditional Food Product of North East India.

A bacteriocin producing strain Lactobacillus brevis UN isolated from Dulliachar-a salted pickle and identified by biochemical and molecular methods. L. brevis UN was found to produce bacteriocin with broad spectrum activity against spoilage causing/food borne pathogens viz. L. monocytogenes, C. perfringens, S. aureus, L. mesenteroides, L. plantarum and B. cereus. Bacteriocin production was optimized through classical one variable at a time method. The isolate showed maximum bacteriocin production at early stationary phase, pH 4.0, temperature 35 °C and with an inoculum size of 1.5 OD @ 10 %. Bacteriocin produced by L. brevis UN was purified to homogeneity by single step gel exclusion chromatography and was most active at pH 6.0 and 7.0, stable up to 100 °C and was proteinaceous in nature. The results of NMR revealed the presence of proline, glutamic acid, aspartic acid, leucine, isoleucine and serine in its peptide structure. PCR amplification analysis determined that bacteriocin encoded gene in L. brevis UN was plasmid bound.

Evaluation of improved functional characteristics of Lactobacillus enriched fruit bar prepared from stone fruits of Mid Himalayan belt.

Probiotic enriched functional fruit bar was prepared by using stone fruits (i.e. apricot and plum) of Mid Himalayan belt. Purpose of this study is to determine the survival of potential probiotic isolates in fruit bar and to study physico-chemical and sensorial properties of the product along with nutritional profile as well as microbial safety. Probiotics bacteria Lactobacillus rhamnosus KC6 and Lactobacillus paraplantarum Sam 1 were used to prepare fruit bar. Probiotic fruit bar was prepared in seven sets and was evaluated for various physico-chemical, nutritional and functional properties during storage period. It was revealed in the study that probiotic fruit bar is nutritionally and functionally better than non-probiotic fruit bar. Probiotic encapsulated bacteria in co-culture combinations efficiently enhanced nutritional quality of fruit bar. Microbiological evaluation of the product revealed that the viability of encapsulated probiotic co-culture bacteria was significantly greater as compared to free cells in the probiotic fruit bar.

Characterization of bacteriocin produced by Brevibacillus laterosporus Tk3 isolated from 'tatwakhar' - a flour prepared from seeds of Aesculus indica in remote areas of Himachal Pradesh India.

Bacteriocin producing strain Brevibacillus laterosporus TK3, was isolated from 'Tatwakhar'- a flour prepared from seeds of Indian Horse Chestnut (Aesculus indica). Bacterial strain Brevibacillus laterosporus TK3 identified by morphological, biochemical techniques followed by 16S rRNA gene sequencing. The 16S rRNA sequence of bacteriocin producer was deposited in NCBI GenBank under accession no. KP861913.1. Bacteriocin of Brevibacillus laterosporus TK3 showed strong antagonistic activity against food spoiling/pathogenic bacteria viz. Listeria monocytogenes, Staphylococcus aureus and Clostridium perfringens. Bacteriocin production by Brevibacillus laterosporus TK3 was enhanced by optimizing production time, pH of the medium, inoculum size and incubation temperature. Maximum bacteriocin activity (6000 AU/ml) was recorded/obtained in basal salt medium of pH 5.5 with an inoculum size of 1.5 OD at 10% and incubation period of 24h at 35 °C. The bacteriocin was purified by single step gel exclusion chromatography. Molecular weight of active bacteriocin from Brevibacillus laterosporous TK3 was found to be 6 kDa according to SDS PAGE. The molecular mass of purified bacteriocin was confirmed as 5953.89 Da by MALDI TOF analysis. The purified bacteriocin was found desirable/suitable for food preservation as it showed wide spectrum of antimicrobial activity, resistance to high temperature, wide pH range and sensitivity to proteolytic enzymes thus, making it safe for human consumption.

Cytology of Rhytidhysteron rufulum: An emerging cause of phaeohyphomycosis.

Fine-needle aspiration cytology (FNAC) is often the first-line investigation for detection of any fungal infection. Rhytidhysteron rufulum is an emerging dematiaceous fungus detected as a human pathogen. FNAC combined with molecular techniques helps in the detection of rare fungal species, especially in cases of non-sporulating fungi. We describe the cytomorphologic features of this species in a 62-year immunocompetent male who presented with a localised subcutaneous infection. Molecular studies helped in the final diagnosis.

Real-time detection of plasma ferritin by electrochemical biosensor developed for biomedical analysis.

Rapid quantification of plasma ferritin levels holds utmost importance for the effective management of different chronic illnesses. We report the development of a novel biosensor for quantitative and selective detection of ferritin from a drop of blood plasma. Developed electrochemical biosensing platform contains a semiconductor nano-structured decorated screen-printed electrode (SND-SPE). The hydrothermally synthesized ZnO-Mn3O4 nanocomposite which has been coated on the electrode surfaces, imparts the specificity in ferritin diagnostics. Cyclic voltametric (CV) measurements with blood plasma shows a prominent reduction peak of ∼ - 0.76 V for specific ferritin reduction. The amperometric sensor shows a known concentration of 0.3 µg/dl ferritin-containing plasma generates 15 µA of current for single-time use. The efficacy of the device is evaluated by detecting ferritin in human plasma samples. The limit of detection and response time of the developed sensor are 0.04 µg/dl and 0.1 s respectively. The layer of ZnO-Mn3O4 nanocomposite has played as an excellent catalyst during the specific reduction of Fe3+ ion and the merits of the device in terms of high robustness, ultrafast detection, highly stable, low-cost, and a biodegradable sensor, make it attractive for the deployment in point-of-care settings.

"Effect of total phenol on the control of leafminer (Phytomyza horticola) infestation in Pea plants".

To determine how vulnerable various pea genotypes are to leafminer infestation, a field experiment was conducted. On the basis of the presence of mines on five randomly selected leaflets from the upper, middle and lower parts of the plant, observations of larvae were made throughout the growing season. The total phenols were determined using the method described by Bray and Thorpe (1954, Analysis of phenolic compounds of interest in metabolism. Methods Biochem Anal. 52:1-27) and absorbance at 650 nm was measured using a spectrophotometer. There was a negative correlation between leafminer infestation and total phenol content. The UHF Pea-12 genotype, characterised by the lowest total phenol concentration (20.87 mg/100 g), exhibited the highest level of leaflet infestation (17.33%). Although UHF Pea-1 genotype had the lowest mean leaflet infestation (6.58%), it also had the highest phenol concentration (41.91 mg per 100 g). In context with this, the present study highlights the significance of host-plant resistance (HPR) in pest management.

Facial chromoblastomycosis in sub-Himalayan region misdiagnosed as cutaneous leishmaniasis: brief report and review of Indian literature.

Cases of chromoblastomycosis, although not uncommon, have rarely been reported in the literature from our region. These are usually misdiagnosed as cases of cutaneous leishmaniasis because we reside in an endemic region for this disease. We present herein patients with facial chromoblastomycosis. They were treated with long-term sodium stibogluconate, but showed no response to therapy. A biopsy revealed these to be cases of chromoblastomycosis, after which the patients were started on antifungal agents. There was marked improvement after the correct treatment was initiated.

Developing health promotional skills among medical undergraduates for tuberculosis patients: An innovative way to benefit students as well as patients.

BACKGROUND: Involving medical undergraduate students in patients' health education will not only help in improving students' learning as well as professional and communication skills but also plays an important role in achieving better health outcome of patients. The aim of the study was to assess the impact of students led educational intervention regarding tuberculosis (TB) on their learning and on the patients' knowledge and attitude towards the disease. MATERIALS AND METHODS: One hundred medical undergraduates were included in the study. An integrated lecture on TB was given to all participants followed by preintervention test on TB. One group was then subjected to interactive traditional lecture. Whereas the videos reflecting the experiences of TB patients were shown to the other group and was given an assignment to prepare educational leaflets for TB patients followed by postintervention test for both groups. The patients' satisfaction level with the information given via educational leaflet and the overall perceived benefits of this activity by the students were assessed. Data analysis done using SPSS version 23.0. Independent and paired t-test were used to calculate difference of means for quantitative variables. RESULTS: Two groups were comparable preintervention however postintervention there was significant improvement in knowledge in the study group (P0.000). Overall 74.2% of the patients were satisfied with the information provided through leaflets. Students' feedback highlighted that majority of the them felt benefitted from this activity. CONCLUSION: It can be concluded that such type of educational interventions have dual benefits, i.e., learning for the students as well as providing health education to patients which in turn will improve their clinical outcome.

Direct N-butyl-2-cyanoacrylate injections to the head and neck for percutaneous embolized devascularization.

BACKGROUND: N-butyl-2-cyanoacrylate (NBCA) has been used for vascular malformations since the 1980s; however, few studies have looked at applications, procedural techniques, and outcome throughout many institutions. Herein, we review applications, procedural techniques, previous literature, and outcomes for the use of NBCA specifically through percutaneous technique in treating head and neck vascular pathology. METHODS: An extensive literature review using PubMed database with published literature containing "N-butyl-2-cyanoacrylate embolization," was performed. No date restrictions were used. Cross-checking of articles was conducted to exclude duplicate articles. The articles were screened for their full text and English language availability. We finalized those articles pertaining to the topic. RESULTS: The search yielded 1124 related articles. When comparing surgical resection to embolization with NBCA for cerebral AVMs, complications were similar in both groups and included hemorrhage (15%), residual AVM (6%), and cerebrospinal fluid leak (3%). Their mortality rate was 3% in both groups. Preoperative percutaneous embolization does show improved surgical outcomes. CONCLUSION: NBCA is a fast-acting liquid embolic material used in the treatment of a variety of vascular malformations and lesions of the head and neck. Investigations surrounding the use of NBCA injections as a new alternative embolic agent began in the 1980's. Administration of NBCA has been shown to be useful in minimizing intraoperative blood loss and controlling acute hemorrhage. Performing percutaneous embolization with NBCA provides a successful alternative for surgeons when transcatheter embolization techniques may prove to be too difficult to perform. Embolization using NBCA will continue to play in integral role in the treatment of malignant lesions and vascular malformations. Continued research is warranted to improve safety, outcomes, and further develop clinical applications of NBCA.

Evaluation of matrix assisted laser desorption ionisation-time of flight mass spectrometry in direct identification of bacteriuria from urine samples.

Objective: The use of matrix-assisted laser desorption/ionisation-time of flight mass spectrometry (MALDI) directly on urine can significantly improve turnaround time for diagnosing urinary tract infection (UTI). The present study was planned to compare the performance of MALDI directly on urine samples with conventional urine culture and evaluate an algorithm using a combination of significant pyuria and MALDI directly on urine samples as a screening method for diagnosing UTI. Materials and Methods: A total of 1000 urine samples from patients with symptoms suggestive of UTIs were analysed. Urine microscopy, urine culture and MALDI were performed simultaneously on all the samples. Results: MALDI correctly identified the microorganisms in 73.83% monomicrobial samples. In culture showing a mixed growth of two and more than three organisms, MALDI was able to identify one microorganism in 27.58% and 15.78% of samples, respectively. There were no peaks by MALDI in 93.53% of 464 sterile samples. The sequential algorithm using urine microscopy and MALDI could correctly identify UTI in 66.23% cases. Conclusion: MALDI can be utilised to rule out bacteriuria in >93% of sterile urine samples. The combination of significant pyuria and direct MALDI as screening method to diagnose UTI (whether monomicrobial or polymicrobial) was not found to be superior than using direct MALDI on urine samples alone.

Nonmissile penetrating spinal injuries: Mechanisms, expectations, and management.

BACKGROUND: Nonmissile penetrating spinal injury (NMPSI) is an uncommon form of traumatic injury to the spine. Here, we present a comprehensive and contemporary literature review that provides insight into NMPSI-type injuries, their mechanisms, clinical practice, management, and expectations. METHODS: An extensive review of the published literature was conducted in PubMed, OVID Medline, and EMBASE journals for studies of nonmissile penetrating spine injuries. Terms for search included NMPSI and nonmissile penetrating spinal cord injury. No date restrictions were used. RESULTS: The search yielded only 17 related articles. Cross-checking of articles was conducted to exclude duplicate articles. The 17 articles were screened for their full text and English language availability. We finalized those articles pertaining to the topic. CONCLUSION: The mechanism of injury in NMPSI occurs in two different stages. Immediate injury is caused by direct damage to the neurological structures. The delayed injury response is caused by damage to the spinal vasculature and activated immune response pathways. Computed tomography (CT) angiograms or formal diagnostic angiograms are indicated to identify vascular injury or development of pseudoaneurysm and can be performed both preoperatively and postoperatively. Surgically, decompressive procedures include laminectomies and hemilaminectomies. Dural exploration may be indicated if a cerebrospinal fluid leak with fistula develops from dural puncture. Further research and technologies are being developed to provide patients who have suffered NMPSI with more resources for a better quality of life.

The Impact of "Vaping" Electronic Cigarettes on Spine Health.

"Vaping" or the use of electronic cigarettes (e-cigarettes) has greatly increased within the past decade, with growing popularity among adolescents. E-cigarettes have many harmful effects on multiple organ systems, but more research is needed to fully understand the extent of possible risks. Our narrative literature review aims to provide comprehensive insight into the impact of e-cigarette use on spinal health with a specific focus on intervertebral disc (IVD) health, bone health, and spinal fusion. There are many metallic compounds and chemical flavoring additives within e-cigarette liquids that are associated with human toxicity. These chemical toxins have been linked to increased oxidative stress leading to systemic inflammation. E-cigarette carcinogens have shown to have a toxic effect on osteoblast cells, and long-term use may decrease bone mineral density and increase the future risk for osteoporosis. Additionally, nicotine in e-liquids negatively impacts IVD health by creating hypoxic environments that degenerate the IVD vasculature and cellular matrix. While studies have demonstrated the inhibitory effects of nicotine use on spinal fusions in animal models, the impact of e-cigarette use on spinal fusion operations in human patients is currently lacking. Future research should focus on the influence of e-cigarette use on spinal health, particularly in adolescents with long-term follow-up, as childhood is a critical time for bone growth and development. Additionally, studies exploring the effects of e-cigarettes on spinal surgery outcomes, such as spinal fusions, are sparse in the literature. Further prospective research studies with a focus on the variety of e-cigarette chemical toxins and flavoring agents is needed to assess the impact on spinal health.

Comparative analysis of virulence determinants, phylogroups, and antibiotic susceptibility patterns of typical versus atypical Enteroaggregative E. coli in India.

Enteroaggregative Escherichia coli (EAEC) is an evolving enteric pathogen that causes acute and chronic diarrhea in developed and industrialized nations in children. EAEC epidemiology and the importance of atypical EAEC (aEAEC) isolation in childhood diarrhea are not well documented in the Indian setting. A comparative analysis was undertaken to evaluate virulence, phylogeny, and antibiotic sensitivity among typical tEAEC versus aEAEC. A total of 171 EAEC isolates were extracted from a broad surveillance sample of diarrheal (N = 1210) and healthy children (N = 550) across North India. Polymerase chain reaction (PCR) for the aggR gene (master regulator gene) was conducted to differentiate tEAEC and aEAEC. For 21 virulence genes, we used multiplex PCR to classify possible virulence factors among these strains. Phylogenetic classes were identified by a multiplex PCR for chuA, yjaA, and a cryptic DNA fragment, TspE4C2. Antibiotic susceptibility was conducted by the disc diffusion method as per CLSI guidelines. EAEC was associated with moderate to severe diarrhea in children. The prevalence of EAEC infection (11.4%) was higher than any other DEC group (p = 0.002). tEAEC occurrence in the diarrheal group was higher than in the control group (p = 0.0001). tEAEC strain harbored more virulence genes than aEAEC. astA, aap, and aggR genes were most frequently found in the EAEC from the diarrheal population. Within tEAEC, this gene combination was present in more than 50% of strains. Also, 75.8% of EAEC strains were multidrug-resistant (MDR). Phylogroup D (43.9%) and B1 (39.4%) were most prevalent in the diarrheal and control group, respectively. Genetic analysis revealed EAEC variability; the comparison of tEAEC and aEAEC allowed us to better understand the EAEC virulence repertoire. Further microbiological and epidemiological research is required to examine the pathogenicity of not only typical but also atypical EAEC.

Purification and characterization of bacteriocin produced by strain of Lactobacillus brevis MTCC 7539.

Bacteriocin, an antimicrobial agent having potential for food biopreservation was purified from Lactobacillus brevis (a safe food-grade bacteria isolated from Vari Kandal, a traditional fermented food of Himachal Pradesh by adopting a novel repeated washing method. Its purity was confirmed by SDS-PAGE and Native-PAGE. The relative molecular mass of bacteriocin was 93.74 kD, while specific activity and recovery were 35.52 folds and 17.13%, respectively. It showed high thermal stability and was active over wide range of pH and exhibited sensitivity to trypsin.

Bacteriocin: safest approach to preserve food products.

Start of the 21st century with its universal call to feed the hungry is an appropriate time to refocus attention on food security and especially the impact of biopatenting on poor communities who are the primary victims of hunger in our world. Antibacterial metabolites of lactic acid bacteria and Bacillus spp have potential as natural preservatives to control the growth of spoilage and pathogenic bacteria in food. Among them, bacteriocin is used as a preservative in food due to its heat stability, wider pH tolerance and its proteolytic activity. Due to thermo stability and pH tolerance it can withstand heat and acidity/alkanity of food during storage condition. Bacteriocin are ribosomally synthesized peptides originally defined as proteinaceous compound affecting growth or viability of closely related organisms. Research is going on extensively to explore the nascent field of biopreservation. Scientists all over the world are showing their keen interest to isolate different types of bacteriocin producing strains and characterize bacteriocin produced by them for food preservation.

Acute Stroke in Tubercular Meningitis: A Case Report.

Most of the strokes in tubercular meningitis are multiple, bilateral, and located in the basal ganglia, especially the 'tubercular zone', which comprises of the caudate, anterior thalamus, anterior limb, and genu of the internal capsule. These are attributed to the involvement of medial striate, thalamotuberal, and thalamostriate arteries, which are embedded in exudates and likely to be stretched by coexistent hydrocephalus. Corticosteroids with antitubercular therapy were thought to reduce mortality and morbidity but their role in lowering strokes has not been proven. The mechanism of stroke in our case was vasculitis. Here, we are reporting a case of 22-years female patient with tubercular meningitis. She had complications of ischemic infarct and severe communicating hydrocephalus with a seizure disorder. Keywords: meningeal tuberculosis; stroke; tuberculoma; vasculitis.

Foetomaternal outcomes of hepatitis E infection outbreak in North India.

Hepatitis E infection (HEV) in pregnant females, especially in the third trimester is associated with poor foetomaternal outcomes. However, the mechanisms of severe liver injury remain obscure. In a recent HEV outbreak in North India, six pregnant females were detected to be positive for HEV infection with concomitant hepatitis A infection in three pregnant females. None of the pregnant females were positive for hepatitis B or hepatitis C infection. The mortality was 50% in pregnant females. In an outbreak, besides, testing for hepatitis markers and understanding the pathogenesis of HEV infection in pregnancy, improving basic hygienic standards is of utmost importance.