A phase II study of ibrutinib in combination with ixazomib in patients with Waldenström macroglobulinaemia.

This phase II study evaluated time-limited (24 cycles) treatment with ibrutinib and ixazomib in newly diagnosed (NDWM; n = 9) and relapsed/refractory (RRWM; n = 12) Waldenström macroglobulinaemia (WM). The overall response rate (ORR) was 76.2% (n = 16) in 21 evaluable patients with no patient achieving a complete response (CR). The median duration of treatment was 15.6 months, and after a median follow-up time of 25.7 months, the median progression-free survival (PFS) was 22.9 months. While the primary end-point was not met (CR rate at any time) and 28.5% discontinued treatment due to toxicity, ibrutinib plus ixazomib led to a clinically meaningful ORR and PFS. Combined Bruton's tyrosine kinase (BTK) and proteasome inhibition merits further evaluation in WM.

Point-of-Care Ultrasound With Artificial Intelligence-Driven Diagnostics in Giant Cell Arteritis: Blindness Prevention on a Global Scale.

Giant cell arteritis (GCA) is the most common autoimmune systemic vasculitis of older adults.1 Typically, it affects individuals 50 years and older, and more women than men. The lifetime risk of GCA for women is 1% and 0.5% for men.

Composition, Treatment, and Outcomes by Radiologically Defined Thrombus Characteristics in Acute Ischemic Stroke.

Occlusive and nonocclusive cervicocephalic thrombi can be encountered during neurovascular imaging in patients with acute ischemic stroke. Radiographic and morphological characteristics on basic and advanced imaging modalities can be important clues towards determination of pathomechanism and the choice of acute and subacute treatment modalities. The aim of this review article is to evaluate the epidemiology, radiographic properties, histologic clot composition of cervicocephalic arterial thrombi, and its response to various medical and endovascular therapy modalities. Future studies are needed to derive and validate a classification system for extracranial and intracranial partially occlusive thrombi to enable further testing of various stroke treatment and prevention strategies in these patients.

Cumulative Effect of pH and Redox Triggers on Highly Adaptive Transient Coacervates.

An intricate synergism between multiple biochemical processes and physical conditions determines the formation and function of various biological self-assemblies. Thus, a complex set of variables dictate the far-from-equilibrium nature of these biological assemblies. Mimicking such systems synthetically is a challenging task. We report multi-stimuli responsive transient coacervation of an aldehyde-appended polymer and a short peptide. The coacervates are formed by the disulphide linkages between the peptide molecules and the imine bond between the polymer and the peptide. Imines are susceptible to pH changes and the disulphide bonds can be tuned by oxidation/reduction processes. Thus, the coacervation is operational only under the combined effect of appropriate pH and oxidative conditions. Taking advantage of these facts, the coacervates are transiently formed under a pH cycle (urea-urease/gluconolactone) and a non-equilibrium redox cycle (TCEP/H2 O2 ). Importantly, the system showed high adaptability toward environmental changes. The transient existence of the coacervates can be generated without any apparent change in size and shape within the same system through the sequential application of the above-mentioned nonequilibrium reaction cycles. Additionally, the coacervation allows for efficient encapsulation/stabilisation of proteins. Thus, the system has the potential to be used for protein/drug delivery purposes in the future.

Skin capillary amylin deposition resembles brain amylin vasculopathy in rats.

BACKGROUND AND PURPOSE: Human amylin is a 37 amino-acid pancreatic peptide that forms neuro-toxic aggregates that deposit in the endothelium of brain capillaries of patients with diabetes, potentially contributing to cerebral small vessel ischemic injury. Pathogenic amylin also deposits in the capillary endothelium in other organs, including the skin. The aim of this study was to test the hypothesis that skin capillary amylin deposition correlates with cerebral small vessel amylin deposition, potentially providing a clinically useful marker of cerebral amylin deposition. METHODS: Immunohistochemistry (IHC) was performed for human amylin and collagen IV in brain and skin sections of rats (age 15-16 months) with pancreatic overexpression of amyloidogenic human amylin polypeptide (HIP rats), and control rats (Wild type; WT; rats that express non-amyloidogenic rat amylin) using antibodies binding amylin (n = 5 male and 5 female rats for each group) and antibodies binding Hypoxia inducing factor (HIF)-1α and HIF-2α (n = 3 for each group). The reactive amylin-aldehyde 4-hydroxynonenal (4-HNE) adduct was measured in skin homogenates. (n = 4 for each group) RESULTS: Brain capillaries isolated from HIP rats had higher amylin content compared to WT rats using Western blot with anti-amylin antibody (p = 0.0010). The HIF-1α and HIF-2α immunoreactivity signals in skin from HIP and WT rats were similar (p = 0.2 for HIF-1 α, and p = 0.75 for HIF-2α). Amylin-4HNE adduct formation was higher in HIP rats compared to WT rats (p = 0.0014). There was phenotypic similarity between brain and skin capillary amylin based on co-staining for human amylin and collagen IV in both HIP and WT rats. CONCLUSION: Skin and brain capillary amylin deposition are similar providing evidence that a skin biopsy might be providing a potential biomarker for diabetes-associated intracranial vasculopathy.

Trends in Racial and Ethnic Diversity in Vascular Neurology Fellowships From 2006 to 2018: A Cross-Sectional Analysis.

BACKGROUND: Workforce diversity in vascular neurology is a crucial component of reducing disparities in stroke care and outcomes. The objective of this study is to describe trends in the racial and ethnic diversity of neurology residents pursuing vascular neurology fellowship and propose an actionable plan for improvement. METHODS: This was a cross-sectional study of race/ethnicity of neurology residents and vascular neurology fellows using published Graduate Medical Education census reports from 2006, when race/ethnicity data were first included, to 2018. Percentage of trainees underrepresented in medicine are reported for 3-year epochs and were analyzed using the Cochran-Armitage test (χ2 test for trend). RESULTS: Across the study period, underrepresented in medicine representation has not changed significantly among all neurology residents and subspecialty fellows (11.9% in 2006-2009; 12.5% in 2015-2018, P=0.82) nor among neurology residents alone (12.0% in 2006-2009; 12.6% in 2015-2018, P=0.81). Among vascular neurology fellows, however, there was a significant downtrend of underrepresented in medicine representation from 16.9% in 2006 to 2009 to 9.3% in 2015 to 2018 (P=0.013). CONCLUSIONS: Racial/ethnic underrepresentation among all neurology residents as well as those pursuing vascular neurology fellowship has persisted across the study period. Concerted efforts should be pursued to increase diversity in neurology residents and vascular neurology fellowship training.

Comparative assessment of multi-trait plant growth-promoting endophytes associated with cultivated and wild Oryza germplasm of Assam, India.

This paper presents a comparative study of endophytic bacteria from cultivated (Oryza sativa) and wild rice (Oryza rufipogon) plants and their functional traits related to plant growth promotion. A total of 70 bacterial isolates were characterized by both biochemical and molecular identification methods. Taxonomic classification showed dominance of three major phyla, viz, Firmicutes (57.1%), Actinobacteria (20.0%) and Proteobacteria (22.8%). Screening for in vitro plant growth-promoting activities revealed a hitherto unreported endophytic bacterium from wild rice germplasm, Microbacterium laevaniformans RS0111 with highest indole acetic acid (28.39 ± 1.39 µg/ml) and gibberellic acid (67.23 ± 1.83 µg/ml) producing efficiency. Few other endophytic isolates from cultivated rice germplasm such as Bacillus tequilensis RHS01 showed highest phosphate solubilizing activity (81.70 ± 1.98 µg/ml), while Microbacterium testaceum MKLS01 and Microbacterium enclense MI03 L05 showed highest potassium (53.42 ± 0.75 µg/ml) and zinc solubilizing activity (157.50%). Fictibacillus aquaticus LP20 05 produced highest siderophore (64.8%). In vivo evaluation of plant growth-promoting efficiencies of the isolates showed that Microbacterium laevaniformans RS0111, Microbacterium testaceum MKLS01 and Bacillus tequilensis RHS 01 could increase rice grain yield by 3.4-fold when compared to the control group. This study indicates the potentiality of rice endophytes isolates as an effective bioinoculants.

A case of aphemia following non-dominant sub-insular stroke: unveiling the Foix-Chavany-Marie phenomenon.

Aphemia refers to the clinical syndrome of inability to orally produce speech with intact comprehension and written expression. Aphemia has been primarily reported in dominant frontal lobe strokes resulting in apraxia of speech (AoS), and in Foix-Chavany-Marie (FCM) syndrome where bilateral opercular or sub-opercular lesions result in anarthria due to deafferentation of brainstem nuclei supplying the oro-facio-lingual and pharyngeal musculature. Aphemia is not reported in non-dominant sub-insular strokes. Here, we present a case of aphemia following non-dominant sub-insular stroke in a patient who had previously recovered from a homologous dominant sub-insular stroke without any apparent residual deficits. We discuss the accepted definitions, theories and controversies in the use of the terminology - aphemia, apraxia of speech (AoS), anarthria related to FCM syndrome, a concomitant pathology - unilateral upper motor neuron (UUMN) dysarthria, and their neuro-anatomical bases. We also highlight the importance of attributing localization value to sequential homologous lesions of the brain that can unveil symptoms due to a "loss of compensation phenomenon" that we propose be termed as "FCM phenomenon." These pathological mechanisms may alone or in certain combinations contribute to the clinical syndrome of aphemia included in the diagnostic approach proposed here. The distinction between these mechanisms requires serial careful neurological examination and detailed speech evaluation including in the recovery phase.

A longitudinal study on morpho-genetic diversity of pathogenic Rhizoctonia solani from sugar beet and dry beans of western Nebraska.

BACKGROUND: Root and stem rot caused by Rhizoctonia solani is a serious fungal disease of sugar beet and dry bean production in Nebraska. Rhizoctonia root rot and crown rot in sugar beet and dry bean have reduced the yield significantly and has also created problems in storage. The objective of this study was to analyze morpho-genetic diversity of 38 Rhizoctonia solani isolates from sugar beet and dry bean fields in western Nebraska collected over 10 years. Morphological features and ISSR-based DNA markers were used to study the morphogenetic diversity. RESULTS: Fungal colonies were morphologically diverse in shapes, aerial hyphae formation, colony, and sclerotia color. Marker analysis using 19 polymorphic ISSR markers showed polymorphic bands ranged from 15 to 28 with molecular weight of 100 bp to 3 kb. Polymorphic loci ranged from 43.26-92.88%. Nei genetic distance within the population ranged from 0.03-0.09 and Shannon diversity index varied from 0.24-0.28. AMOVA analysis based on ΦPT values showed 87% variation within and 13% among the population with statistical significance (p < 0.05). Majority of the isolates from sugar beet showed nearby association within the population. A significant number of isolates showed similarity with isolates of both the crops suggesting their broad pathogenicity. Isolates were grouped into three different clusters in UPGMA based cluster analysis using marker information. Interestingly, there was no geographical correlation among the isolates. Principal component analysis showed randomized distribution of isolates from the same geographical origin. Identities of the isolates were confirmed by both ITS-rDNA sequences and pathogenicity tests. CONCLUSION: Identification and categorization of the pathogen will be helpful in designing integrated disease management guidelines for sugar beet and dry beans of mid western America.

Smart Thixotropic Hydrogels by Disulfide-Linked Short Peptides for Effective Three-Dimensional Cell Proliferation.

Supramolecular assembly of short peptides is a crucial process and has shown numerous potential applications as biomaterials. In the present work, the hydrogelation process of short peptides containing C-terminal "Lys-Cys" (KC) residues have been studied in detail. The N-terminal capping is found to be essential for effective gelation. Out of 12 peptides we studied, two of them could form hydrogels efficiently: Ac-VVKC-NH2 and Ac-FFKC-NH2. In both cases, the monomer-to-dimer formation through disulfide linkages by Cys residues controls the aggregation process. Interestingly, the presence of H2O2 facilitated the dimerization and thereby reduced the gelation time but could not impart much effect on the mechanical properties of the gels. Detailed rheological study revealed that both hydrogels are thixotropic in nature. Moreover, they are responsive to glutathione (GSH) due to the presence of disulfide linkages. However, the hydrogel of Ac-FFKC-NH2 is found to be stronger and more effective for biological applications. The thixotropic nature as well as a model drug release study in response to varying GSH concentration indicates the possible use of the hydrogel as an injectable local drug delivery vehicle. The hydrogel of Ac-FFKC-NH2 is noncytotoxic in nature. Three-dimensional cell proliferation has been found to be more effective than 2D, as it mimics the in vivo situation more closely if not exactly. In the present study, we have shown that both differentiated RAW macrophages and undifferentiated THP-1 monocytes could proliferate significantly within the 3D matrix of the hydrogel, without depicting any apparent cytotoxicity. Thus, the hydrogel of Ac-FFKC-NH2 has potential for application in localized drug administration and as a supporting biomaterial to study basic phenomena involving cell behavior.

Characterization of siderophore producing arsenic-resistant Staphylococcus sp. strain TA6 isolated from contaminated groundwater of Jorhat, Assam and its possible role in arsenic geocycle.

BACKGROUND: Microorganisms specifically bacteria play a crucial role in arsenic mobilization and its distribution in aquatic systems. Although bacteria are well known for their active participation in the different biogeochemical cycles, the role of these bacteria in regulating the concentration of arsenic in Brahmaputra valley has not been investigated in detail. RESULTS: In this paper, we report the isolation of an arsenic resistant bacterium TA6 which can efficiently reduce arsenate. The isolate identified as Staphylococcus sp. TA6 based on the molecular and chemotaxonomic identification (FAME) showed resistance to the high concentration of both arsenate and arsenite (As(III) = 30 mM; As(V) = 250 mM), along with cross-tolerance to other heavy metals viz., Hg2+, Cd2+, Co2+, Ni2+, Cr2+. The bacterium also had a high siderophore activity (78.7 ± 0.004 µmol) that positively correlated with its ability to resist arsenic. The isolate, Staphylococcus sp. TA6 displayed high bio-transformation ability and reduced 2 mM As(V) initially added into As(III) in a period of 72 h with 88.2% efficiency. The characterization of arsenate reductase enzyme with NADPH coupled assay showed the highest activity at pH 5.5 and temperature of 50 °C. CONCLUSIONS: This study demonstrates the role of an isolate, Staphylococcus sp. TA6, in the biotransformation of arsenate to arsenite. The presence of ars operon along with the high activity of the arsenate reductase and siderophore production in this isolate may have played an important role in mobilizing arsenate to arsenite and thus increasing the toxicity of arsenic in the aquatic systems of the Brahmaputra valley.

Is 400 IU per day of Vitamin-D given to healthy well-nourished mothers antenatally enough to prevent neonatal Vitamin-D deficiency?

BACKGROUND: The incidence of Vitamin D deficiency (VDD) among newborns is very high in India, this is usually secondary to maternal deficiency. Currently Vitamin D at the dose of 400-600 IU/day is being prescribed during pregnancy for prevention. There is paucity of data regarding efficacy of current recommendations in preventing neonatal hypovitaminosis D.Our aim was to determine if good nutritional status of the mother and regular calcium and Vitamin D supplementation during pregnancy in the currently recommended doses help prevent or reduce VDD in neonates. METHODS: Cord blood Serum 25(OH)D levels were studied in 342 neonates born to 323 healthy mothers at a tertiary care center in Mumbai. All the mothers were on regular antenatal follow up and received Vitamin D and calcium supplements as per current recommendations. RESULTS: Mean umbilical cord blood 25(OH)D level was 12.088 ng/ml (SD 3.349, range 31.1-5.1). Out of 342 neonates, 335 (97.95%) had Vitamin D deficiency [cord blood 25(OH)D level <20 mg/ml] 1.75% had cord blood 25(OH)D level between 21 and 29 ng/ml in the insufficiency range and only 0.29% of the subjects had 'normal' cord blood 25(OH)D levels of >30 ng/ml. However, none of the neonates in the study were found to have severe VDD (<5 ng/ml). CONCLUSION: Antenatal supplementation of well nourished, socioeconomically better off mothers in our population with 400 IU/day of Vitamin D3 is ineffective in preventing VDD in neonates.

Nitrogen dynamics as a function of soil types, compaction, and moisture.

In this study, the complex interactions between soil types, compaction, and moisture on nitrogen (N) transformation processes such as ammonia (NH3) volatilization, ammonification, nitrification, and denitrification were examined over a 30-day period using a simulated column approach. Two soil types: loam, and sandy loam, were subjected to three compaction treatments-control, surface, and sub-surface compaction-and two moisture regimes, dry and wet. Liquid urea ammonium nitrate (32-0-0) was used as the N fertilizer source at a rate of 200 kg N ha-1. Key indicators of N transformations were measured, including residual concentrations of ammonium (NH4-N) and nitrate (NO3-N), NO3-N leaching, NH3 volatilization, and nitrous oxide (N2O) emissions. Findings revealed that compaction significantly increased residual NH4-N concentrations in deeper soil profiles, with the highest 190.80 mg kg-1 recorded in loam soil under sub-surface compaction and dry conditions. Nitrification rates decreased across both soil types due to compaction, evidenced by elevated residual NH4-N levels. Increased NO3-N leaching was observed in loam soil (178.06 mg L-1), greater than sandy loam (81.11 mg L-1), due to initial higher residual NO3- in loam soil. The interaction of compaction and moisture most affected N2O emissions, with the highest emissions in control treatments during dry weather at 2.88 kg ha -1. Additionally, higher NH3 volatilization was noted in moist sandy loam soil under control conditions at 19.64 kg ha -1. These results highlight the necessity of considering soil texture, moisture, and compaction in implementing sustainable N management strategies in agriculture and suggest recommendations such as avoiding broadcast application in moist sandy loam and loam soil to mitigate NH3 volatilization and enhance N use efficiency, as well as advocating for readjustment of fertilizer rate based on organic matter content to reduce potential NO3-N leaching and N2O emissions, particularly in loam soil.

Factors determining utilization of stem cell transplant for initial therapy of multiple myeloma by patient race: exploring intra-racial healthcare disparities.

Multiple myeloma (MM) therapeutics have evolved tremendously in recent years, with significant improvement in patient outcomes. As newer treatment options are developed, stem cell transplant (SCT) remains an important modality that provides excellent disease control and delays the progression of disease. Over the years, SCT use has increased overall in the U.S., but two distinct gaps remain, including suboptimal use overall and racial-ethnic disparities. We evaluated the National Cancer Database (NCDB) to study what sociodemographic factors might play a role within a given racial-ethnic group leading to disparate SCT utilization, such that targeted approaches can be developed to optimize SCT use for all. In nearly 112,000 cases belonging to mutually exclusive categories of non-Hispanic Whites (NHW), non-Hispanic Blacks (NHB), Hispanics, non-Hispanic Asians (NHA), and others, we found certain factors including age, comorbidity index, payor type, facility type (academic vs. community) and facility volume to be uniformly associated with SCT use for all the racial-ethnic groups, while gender was not significant for any of the groups. There were several other factors that had a differential impact on SCT utilization among the various race-ethnicity groups studied, including year of diagnosis (significant for NHW, NHB, and Hispanics), income level (significant for NHW and Hispanics), literacy level (significant for NHW and NHB), and geographic location of the treatment facility (significant for NHW and NHA). The suboptimal SCT utilization overall in the U.S. suggests that there may be room for improvement for all, even including the majority NHW, while we continue to work on factors that lead to disparities for the traditionally underserved populations. This study helps identify sociodemographic factors that may play a role specifically in each group and paves the way to devise targeted solutions such that resource utilization and impact can be maximized.

Anti-B Cell Maturation Antigen Chimeric Antigen Receptor T Cell Therapy for the Treatment of AL Amyloidosis and Concurrent Relapsed/Refractory Multiple Myeloma: Preliminary Efficacy and Safety.

While immunotherapies, such as CAR T therapy and bi-specific antibodies, have revolutionized the treatment of multiple myeloma (MM), patients with AL amyloidosis have been excluded from trials with these agents due to concerns of underlying autonomic, cardiac, and renal dysfunction, leading to potentially fatal toxicities from these therapies. In this communication, we described the outcomes of two patients with AL amyloidosis and concurrent MM with underlying cardiac and/or renal dysfunction who underwent anti-BCMA CAR T cell therapy with ide-cel or cilta-cel, received cytokine release syndrome prophylaxis, and tolerated therapy well with manageable toxicities and achieved a MRD-negative state. We described the preliminary efficacy and safety of CAR T in patients with AL amyloidosis and highlighted the importance of patient selection and medical optimization of cardiac and renal function prior to CAR T.

Evaluation of kit-based loop-mediated isothermal amplification (TB-LAMP) assay in the diagnosis of tubercular lymphadenitis.

Background: The rapid and accurate diagnosis of tubercular lymphadenitis remains a challenging task today. The World Health Organization (WHO) endorsed the LoopAMP MTBC kit (TB-LAMP) as a replacement for sputum smear microscopy in the diagnosis of pulmonary tuberculosis (PTB). However, no prospective diagnostic accuracy study of TB-LAMP for tubercular lymphadenitis in adults has been performed yet. The current study evaluated the diagnostic performance of TB-LAMP in tubercular lymphadenitis (LNTB). Methods: In a prospective observational study conducted at a tertiary care hospital in India, 90 subjects (age >18 years) suspected of LNTB were recruited consecutively and followed up for 6 months between January 2019 and December 2020. Samples were processed for microscopy, culture, GeneXpert, histopathology and TB-LAMP. The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of TB-LAMP against the composite reference standard (CRS) and culture were determined. Results: TB-LAMP showed a sensitivity of 83.78 % (95 % CI, 73.76-90.47) and a specificity of 81.25 % (95 % CI, 56.99-93.41), respectively, against the CRS. The PPV and NPV were 95.38 % (95 % CI, 87.29-98.42) and 52.00 % (95 % CI, 33.50-69.97), respectively. TB-LAMP showed a sensitivity of 88.89 % (95 % CI, 71.94-96.15) and a specificity of 36.17 % (95 % CI, 23.97-50.46), respectively, against culture. The PPV and NPV were 44.44 % (95 % CI, 32-57.62) and 85 % (95 % CI, 63.96-94.76), respectively. Conclusion: TB-LAMP can be used instead of conventional microscopy for the diagnosis of TB in lymph node specimens at primary healthcare centres. It provides rapid and cost-effective diagnosis of LNTB in resource-limited settings due to good sensitivity and NPV.

The digital divide: Racial disparities in adoption and utilization of health information technology among patients with lymphoid cancers.

INTRODUCTION: Health information technology (HIT) has the potential to improve healthcare delivery and engagement. Studying racial-ethnic disparities in HIT engagement will help understand and overcome challenges to healthcare utilization. METHODS: We undertook a patient-reported survey among patients with lymphoid malignancies at two campuses of Mayo Clinic, Florida to explore HIT-related disparities. Variables between Whites and non-Whites, and non-Whites from the two campuses were compared. RESULTS: The survey was completed by 1004 respondents, with 71% whites, 27% non-Whites (race-ethnicity not reported by 2%). Non-Whites included 30% responders at the main campus and 64% at an inner-city campus. Whites were significantly older and had higher education, while non-Whites had lesser access to a computer. Only 51% of non-Whites were registered to use electronic medical records (EMR) as compared to 72% Whites (p < 0.001) and significantly lesser number of non-Whites even knew that EMR existed (81% vs. 92%, p < 0.001). Encouragingly, a higher number of non-Whites wanted to engage in EMR. Non-Whites from the main campus were older, more educated and had more access to a computer as compared to those from the inner-city campus. Similar disparate factors were noted among minorities from the two campuses, suggesting impact of socioeconomic backgrounds on EMR usage among non-Whites. Linguistic barriers were more striking among inner-city campus non-Whites. CONCLUSIONS: Non-Whites continue to struggle with suboptimal utilization of the healthcare system and barriers related to integration in HIT, including disparities representing socioeconomic differences. Efforts need to be made at several levels to help racial-ethnic minorities overcome awareness, access, and linguistic barriers to HIT utilization.

Microliter spotting and micro-colony observation: A rapid and simple approach for counting bacterial colony forming units.

For enumerating viable bacteria, traditional dilution plating to count colony forming units (CFUs) has always been the preferred method in microbiology owing to its simplicity, albeit being laborious and time-consuming. Similar CFU counts can be obtained by quantifying growing micro-colonies in conjunction with the benefits of a microscope. Here, we employed a simple method of five to ten microliter spotting of a diluted bacterial culture multiple times on a single Petri dish followed by determining CFU by counting micro-colonies using a phase-contrast microscope. In this method, the CFU of an Escherichia coli culture can be estimated within a four-hour period after spotting. Further, within a ten-hour period after spotting, CFU in a culture of Ralstonia solanacearum, a bacterium with a generation time of around 2 h, can be estimated. The CFU number determined by micro-colonies observed for 106-fold dilutions or lower is similar to that obtained by the dilution plating method for 107-fold dilutions or lower. Micro-colony numbers observed in the early hours of growth (2 h in case of E. coli and 8 h in case of R. solanacearum) were found to remain consistent at later hours (4 h in case of E. coli and 10 h in case of R. solanacearum), where the visibility of the colonies was better due to a noticeable increase in the size of the colonies. This suggested that micro-colonies observed in the early hours indeed represent the bacterial number in the culture. Practical applications to this counting method were employed in studying the rifampicin-resistant mutation rate as well as performing a fluctuation test in E. coli. The spotting method described here to enumerate bacterial CFU results in reduction of labour, time and resources.