Characterization, histopathology and immunogenicity of the lumpy skin disease virus isolated during 2019-20 in Bangladesh.

Introduction: Lumpy skin disease (LSD) is a highly contagious vector-borne viral disease of cattle. LSD has emerged in Bangladesh in 2019, causing significant economic losses due to its high morbidity and mortality. This research was designed to isolate, identify, and assess the immunogenicity of LSD virus (LSDV) using nodular tissue samples obtained from affected cattle during the 2019-20 outbreak across nine districts of Bangladesh. Methods: To determine the presence of LSDV in nodular tissues, we initially used iiPCR and PCR, followed by histopathological examination. 151 were positive via iiPCR and PCR among the 180 collected samples. The PCR positive 151 samples were then inoculated into 10-day-old embryonated chicken eggs via the CAM route to isolate LSDV, confirmed through PCR. Subsequently, partial sequencing and phylogenetic analysis of the P32 gene were performed to determine the origin of the circulating LSDV strain. The immunogenicity of selected LSDV strains was assessed through an ELISA test. Results: The PCR results revealed a distinct positive band at 192 bp in both the nodular tissue samples and the LSDV isolated from chicken embryo inoculations. Microscopic analysis of the nodular lesions revealed thickening of the epidermis, ballooning degeneration of keratinocytes, and proliferation of follicular epithelia. Additionally, mononuclear infiltration was observed at the demarcation line between infected and healthy tissue, with necrosis of muscular tissues beneath the epidermis. The LSDV isolate from Bangladesh exhibited a close genetic relationship with LSDV strains isolated from neighboring and other regional countries including India, Myanmar, and Mongolia. This observation strongly suggests the possibility of a transboundary spread of the LSD outbreak in Bangladesh during 2019-2020. The results of the immunogenicity test showed that the serum antibody titer remained at a protective level for up to 18 months following secondary immunization with inactivated LSDV antigen. This finding suggests that the inactivated LSDV antigen could be a potential vaccine candidate to protect cattle in Bangladesh against LSDV. Conclusion: In conclusion, our research successfully isolated, identified, and characterized LSDV in cattle nodular tissues from the 2019-20 outbreak in Bangladesh. Furthermore, it provided insights into the probable origin of the circulating strain and investigated a potential vaccine candidate to protect cattle in the region from LSDV.

Complete genome sequence analysis of the multidrug resistant Aeromonas veronii isolated for the first time from stinging catfish (Shing fish) in Bangladesh.

Objective: Whole genome sequencing (WGS) of Aeromonas veronii Alim_AV_1000 isolated from ulcerative lesions of Shing fish (stringing catfish; Heteropneustes fossilis) was performed during the outbreak year 2021. Materials and Methods: Using next-generation sequencing (Illumina) technology, WGS was accomplished, resulting in the sequencing, assembly, and analysis of the entire genome of the A. veronii strain. Moreover, the genomic features, virulence factors, antimicrobial resistome, and phylogenetic analysis for the molecular evolution of this strain were also examined. Results: The genome size of the A. veronii Alim_AV_1000 strain was 4,494,515 bp, with an average G+C content of 58.87%. Annotation revealed the known transporters and genes linked to virulence, drug targets, and antimicrobial resistance. Conclusion: The findings of the phylogenetic analysis revealed that the strain of the present study has a close relationship with the China strain TH0426 and strain B56. This study provides novel information on A. veronii isolated from Shing fish in Bangladesh.

Effect of knowledge of informal poultry drug prescribers on their attitude and practice toward antimicrobial use, residues, and resistance in Bangladesh.

Background and Aim: Informal prescribers (IPs) significantly contribute to the development of antimicrobial resistance and in disseminating pathogens from poultry to humans and other animals through the food chain, posing a serious global health threat. Therefore, this study aimed to assess whether the knowledge of IPs has an impact on their attitude and practice toward antimicrobial use, antibiotic residues, and antimicrobial resistance. Materials and Methods: In this cross-sectional study, we conducted a pre-tested and questionnaire-based survey to investigate the knowledge, attitude, and practice of IPs in selected parts of the Mymensingh division, Bangladesh. Then, we used the linear regression model test with R-squared (R2) to measure the association between the study variables. Results: Our investigation revealed that 70% of the IPs knew about antibiotics and 75% had good knowledge about antibiotic resistance, whereas only 50% were aware of withdrawal periods. Informal prescribers also displayed good attitudes toward the use and sale of antibiotics with withdrawal periods and completion of medication (50%). Analysis of their practice on the sale and prescription of antibiotics showed that 70% and 30% of IPs use antibiotics against bacterial infections and other conditions, respectively. Most of them do not consult a veterinarian before selling or prescribing antibiotics, although 80% claim to do so. This is because 75% of IPs gave other options regarding their consultations. However, 95% of IPs uses antibiotics only for therapeutic purposes. Furthermore, only 10% sell antibiotics based on a veterinarian's recommendation. Approximately 45% of IPs use single antibiotics at a time, while the rest use multiple antibiotics, individually or combined. Approximately 15% use antibiotics monthly, while 85% use them whenever the need arises. The knowledge and attitude of IPs are significantly affected by their age (p ≤ 0.025). The district of domicile also impacted their knowledge. Surprisingly, IPs from Jamalpur had significantly better knowledge compared to those from Mymensingh and Sherpur (p ≤ 0.01). The attitude of IPs from Jamalpur and Netrokona also differed significantly (p ≤ 0.001) from that of Mymensingh and Sherpur. The knowledge of IPs influenced their attitude up to 80.5% (r2 = 0.628) and their practice up to 75.4% (r2 = 0.545). Conclusion: The knowledge of IPs greatly influenced their attitude and practice, while sociodemographics also influenced their knowledge and attitude toward antimicrobial use, antibiotic residues, and antimicrobial resistance.

Facile synthesis of CuONPs using Citrus limon juice for enhancing antibacterial activity against methicillin-resistant Staphylococcus aureus, beta-lactamase and tetracycline-resistant Escherichia coli.

Antimicrobial resistance (AMR) resulting from indiscriminate use of antibiotics in various fields of agriculture such as livestock farming, aquaculture, and croup fields become an emerging catatroph for the health (human, animal) and environment. Among those, poultry farming has been considered as one of the major contributors of multidrug-resistant (MDR) bacteria. Focusing this, the present research is designed for green synthesis of copper oxide nanoparticles (CuONPs) with the aim of their application in antibiotic-free poultry farming for curving use of antibiotics in that sector. For that, antibacterial CuONPs were nanoformulated to decrease the required doses of bulk CuSO4. We used a CuSO4·5H2O solution as a Cu2+ source and Citrus limon juice as a reducing agent as well as capping agent. Particle yield was initially confirmed by the λmax specific to CuONPs (295 nm) using UV-Vis spectroscopy. The presence of the Cu-O group during particle formation and crystallinity with the purity of yielded NPs was confirmed with Fourier-transform infrared spectroscopy and X-ray diffractometry. The round to spherical CuONPs of 92-155 nm average size was confirmed with atomic force, scanning electron, and transmission electron microscopy. The concentration of yielded NPs was calculated with the dynamic light scattering. The physical characterization tools indicated a maximum CuONPs yield with a 0.001 M ion source with 15% reducing agents after 12 h reduction. Antibacterial effectivity was tested against methicillin-resistant Staphylococcus aureus and tetracycline- and beta-lactamase-resistant Escherichia coli, confirmed by PCR amplicon band at 163 bp, 643 bp, and 577 bp for the mecA, blaTEM-1 and tetA genes, respectively. An antibiogram assay of CuONPs showed a maximum zone of inhibition of 26 ± 0.5 mm for the synthesized particles. The minimum inhibitory and bactericidal concentrations were 1.6 µg ml-1 and 3.1 µg ml-1, respectively, for broad-spectrum application. Finally, the biocompatibility of CuONPs was determined by demonstrating a nonsignificant decrease of BHK-21 cell viability at <2 MIC doses for complying their future in vivo applicability.

Antimicrobial Resistance Profile of Common Foodborne Pathogens Recovered from Livestock and Poultry in Bangladesh.

Multidrug-resistant (MDR) foodborne pathogens have created a great challenge to the supply and consumption of safe & healthy animal-source foods. The study was conducted to identify the common foodborne pathogens from animal-source foods & by-products with their antimicrobial drug susceptibility and resistance gene profile. The common foodborne pathogens Escherichia coli (E. coli), Salmonella, Streptococcus, Staphylococcus, and Campylobacter species were identified in livestock and poultry food products. The prevalence of foodborne pathogens was found higher in poultry food & by-product compared with livestock (p < 0.05). The antimicrobial drug susceptibility results revealed decreased susceptibility to penicillin, ampicillin, amoxicillin, levofloxacin, ciprofloxacin, tetracycline, neomycin, streptomycin, and sulfamethoxazole-trimethoprim whilst gentamicin was found comparatively more sensitive. Regardless of sources, the overall MDR pattern of E. coli, Salmonella, Staphylococcus, and Streptococcus were found to be 88.33%, 75%, 95%, and 100%, respectively. The genotypic resistance showed a prevalence of blaTEM, blaSHV, blaCMY, tetA, tetB, sul1, aadA1, aac(3)-IV, and ereA resistance genes. The phenotype and genotype resistance patterns of isolated pathogens from livestock and poultry had harmony and good concordance, and sul1 & tetA resistance genes had a higher prevalence. Good agricultural practices along with proper biosecurity may reduce the rampant use of antimicrobial drugs. In addition, proper handling, processing, storage, and transportation of foods may decline the spread of MDR foodborne pathogens in the food chain.

Epidemiological investigation of lumpy skin disease outbreaks in Bangladeshi cattle during 2019-2020.

Lumpy skin disease (LSD) is an acute infectious viral disease of cattle with a wide distribution that emerged in Bangladesh in 2019, causing huge economic losses. This study was undertaken to investigate the epidemiological features of LSD emergence in nine districts of Bangladesh between December 2019 and December 2020. A total of 8215 cattle from 603 herds were investigated and LSD was diagnosed based on the characteristic clinical findings. A standard questionnaire was administered to collect herd-level data including location, herd size, number of LSD-infected cattle, number died due to LSD, farm type, season, house type, vector presence, sanitation and fly repellent use. Similarly, data on clinical signs, sex, age, animal class and breed of the LSD-infected cattle were also recorded. The herd-level attack risk (%) and mortality risk (%) were calculated based on the number of infected and dead cattle, respectively, as a proportion of total cattle. The herd-level risk factors for LSD were identified using a multivariable Poisson regression model. The most common clinical signs were skin nodules (100%), fever (97.9%) and depression with anorexia and weight loss (97.9%). Crossbred (84.9%) and female (72.2%) cattle were mostly affected by LSD. The overall LSD attack risk, mortality risk and case fatality were 26.5%, 0.26% and 0.97%, respectively. The LSD attack risk was significantly higher in small herds (risk ratio: [RR] 1.39; 95% CI: 1.27; 1.53) than large herds. In addition, significantly higher LSD attack risk was observed in semi-intensive management systems (RR = 1.29; 95% CI: 1.01; 1.64) than intensive management systems. Moreover, it was also significantly higher in hut (RR = 1.81; 95% CI: 1.12; 2.92), temporary (RR = 1.62; 95% CI: 1.21; 2.17) and tin-shed houses (RR = 1.29; 95% CI: 1.11; 1.51) than in semi-building houses. To the best of our knowledge, this is the first detailed epidemiological study of LSD emergence in South Asia. Female crossbred cattle in small herds under semi-intensive management should be prioritized for LSD surveillance and vaccination to prevent further outbreaks and control the impact of the disease in Bangladesh.

Multi-species probiotics improve growth, intestinal microbiota and morphology of Indian major carp mrigal Cirrhinus cirrhosus.

This study aimed to examine the effects of multi-species probiotic on growth, hematological status, intestinal microbes, and intestinal morphology of mrigal (Cirrhinus cirrhosus). The mrigal fries (average weight 0.51 g) were reared for 60 days by supplementing multi-species probiotics containing Bacillus spp. (1 × 109 cfu/mL) and Lactobacillus spp. (1 × 1011 cfu/mL) in the raising water at doses of 0 (control), 0.5, and 1.0 mL/L. The results indicated that fish reared with multi-species probiotics showed significantly higher growth performance and feed efficiency where the survival rate was similar in all cases. Accordingly, significant higher red blood cell (RBC) and white blood cell (WBC) were counted from the fish reared with multi-species probiotic. There was a considerable difference in bacterial microbiota between the experimental and control group. Multi-species probiotics significantly enhanced the length, width, and villus area. Several immune response indicators like fattening of intestinal mucosal fold, width of lamina propria, the width of enterocytes, and abundance of goblet cells were also increased significantly in fish that received multi-species probiotics. This study revealed that multi-species probiotics can significantly contribute to the growth of mrigal through upgrading intestinal microbiota and morphology, which can be suggested as an eco-friendly growth stimulator in mrigal farming.

Efficacy of bi-valent whole cell inactivated bacterial vaccine against Motile Aeromonas Septicemia (MAS) in cultured catfishes (Heteropneustes fossilis, Clarias batrachus and pangasius pangasius) in Bangladesh.

The Motile Aeromonas Septicemia (MAS) is an important disease of cultured catfishes (Heteropneustes fossilis, Clarias batrachus and Pangasius pangasius), caused by different species of Aeromonas bacteria which have been documented to be higher death rates (≤70%) in Bangladesh since 2016. Present study was conducted to develop bi-valent vaccine using A. hydrophila and A. veronii, and to validate their efficacy via intra-muscular (IM) and oral-routes of immunization in selected species of fishes. Brood fishes of the three species were immunized with three doses of inactivated vaccine (107 CFU /2.3 mg/ml). Hematological parameters of brood fishes and antibody levels (IgM) of broods, their larvae and eggs were determined by ELISA. Additionally, Relative Percent Survivability (RPS) and the IgM levels of the larvae after challenge with virulent A. hydrophila and A. veronii were also evaluated. Findings of this study showed that the lymphocytes, monocytes, granulocytes counts and antibody (IgM) titre of brood fishes, larvae and eggs from the vaccinated fishes were found significantly higher (p< 0.05) compared to the un-vaccinated control groups. Alternatively, antibody levels (IgM) in the larvae of vaccinated group of brood fishes fed with antigen coated feed was exhibited to be remarkably higher (p< 0.05) than the antigen non-fed group. The RPS of larvae of Shing (91.24 ± 2.00%), Magur (88.09 ± 2.88%) and Pangas (93.17 ± 1.52%) was found to be higher in the larvae at 20-day age of vaccinated group compared to non-vaccinated brood fishes group. Findings of this study indicated that the active immunization of brood fishes followed by oral immunization of their larvae feeding with antigen coated feed showed synergistic effect in protecting cultured Shing, Magur and Pangas fishes from frequent attack with Aeromonas spp at any age of their lifetime.

A Survey on Knowledge, Attitude, and Practices of Large-Animal Farmers towards Antimicrobial Use, Resistance, and Residues in Mymensingh Division of Bangladesh.

The widespread and indiscriminate use of antimicrobials in food animals is a key contributor to antimicrobial resistance and antimicrobial residue, which have become a growing public and animal health concern in developing countries such as Bangladesh. This study was aimed to assess the knowledge, attitude, and practices (KAP) of large-animal farmers towards antimicrobial use (AMU), antimicrobial resistance (AMR), and antimicrobial residue (AR) with their correlation. A cross-sectional survey was conducted with a structured and pretested questionnaire in the Mymensingh division of Bangladesh. A total of 212 large-animal farmers (dairy, beef fattening, buffalo, sheep, and goat farmers) were surveyed. Results showed that most of the farmers are male (85.8%) and belong to the 18-30 age group (37.3%). About 20.3% had no formal education, and nearly half of the participants (48.1%) received training regarding antibiotic use and resistance. Penicillin is the most common class of antibiotic used (61.8%) in the study area, followed by other antimicrobials. Only 37.7% of the farmers used antimicrobials on the recommendation of their veterinarian. Overall, 41.5%, 42.5%, and 21.7% of farmers possess adequate knowledge and a satisfactory attitude and perform desirable practices, respectively. Farmers in the 31-40 age group have adequate knowledge, attitude, and ability to implement desired practices compared to farmers in the 18-30 age group. Farmers having a graduate or post-graduate degree scored better in relation to knowledge, attitude, and practice than other farmers. Analysis revealed that farmers who received training on AMU and AMR had 10.014 times (OR = 10.014, 95% CIs: 5.252-19.094), 9.409 times (OR = 9.409, 95% CIs: 4.972-17.806), and 25.994 times (OR = 25.994, 95% CIs: 7.73-87.414) better knowledge, attitude, and performance, respectively, compared to their counterparts. A significant proportion of farmers (97.2%) dispose of leftover antibiotics inappropriately. The findings of the present study will be used to intervene in the education and training of the farmers, which will help to limit the indiscriminate and irrational use of antimicrobials, leading to reducing the chances of developing AMR.

groEL gene-based molecular detection and antibiogram profile of Riemerella anatipestifer from duck in Bangladesh.

Objectives: This study was designed to detect Riemerella anatipestifer through polymerase chain reaction (PCR) from duck farming areas of the Mymensingh and Sylhet divisions and to determine the antibiogram profile of the PCR-positive isolates using the disc diffusion method. Materials and Methods: Fifty two samples were collected, comprising clinically sick (32 ducks) and dead ducks (20). PCR confirmation was accomplished, and consistent findings were observed, employing R. anatipestifer groEL (271-bp) gene as appropriate molecular markers. For further clarification, see R. anatipestifer specific PCR assay (546-bp) and gyrB-based PCR (162-bp) were also done. The disc diffusion method was followed for the antibiotic susceptibility test of the isolates against commonly used antibiotics. Results: A total of 21 samples, 8 from clinically sick birds and 13 from dead birds, showed positive results in both conventional and molecular assays out of 52 samples. High occurrences were found in oropharyngeal swabs from sick ducks and the liver and heart from dead ducks. Antibiotic susceptibility testing revealed that the isolates were 100% resistant to penicillin G, cefradine, streptomycin, neomycin, gentamycin, meropenem, and erythromycin, but 100% sensitive to -cotrimoxazole, florfenicol, and levofloxacin. Conclusion: For diverse duck-populated areas in Bangladesh, this study shows the severity of R. anatipestifer infection among ducks.

Isolation of multidrug-resistant Escherichia coli and Salmonella spp. from sulfonamide-treated diarrheic calves.

Background and Aim: The bovine industry is threatened by one of the most serious and deadly enteric diseases, calf diarrhea, particularly in developing nations like Bangladesh. In this context, bacterial resistance to antimicrobial drugs and its detrimental consequences have become a critical public health issue that is difficult to address globally. This study aimed to isolate and identify Escherichia coli and Salmonella spp. with their antibiogram and antibiotic resistance gene detection from sulfonamide-treated diarrheic calves. Materials and Methods: Twelve diarrheic calves suffering from calf diarrhea in a dairy farm were selected and a total of 36 fecal samples were aseptically collected directly from rectum before, during, and at the end of treatment for each calf to determine the total viable count, total E. coli count and total Salmonella count. A polymerase chain reaction was used for the specific detection of E. coli and Salmonella genus targeting fliC and invA genes, respectively. Antibiotic sensitivity test of the isolated E. coli and Salmonella spp. were performed by the disk diffusion method for eight commonly used antibiotics. Results: A total of 36 E. coli (100%) and 12 Salmonella spp. (33%) were isolated from the samples and were confirmed by polymerase chain reaction. Total viable count was found to be ranged from 35 × 107 to 99 × 1010 colony-forming unit (CFU)/g fecal sample before starting sulfonamide treatment, 34 × 105 to 25 × 1010 CFU/g during treatment with sulfonamide, and 48 × 103 to 69 × 1010 CFU/g immediately after completion of sulfonamide treatment. Total E. coli count was found to be ranged from 4 × 104 to 36 × 1010 CFU/g, 24 × 104 to 23 × 108 CFU/g, and 13 × 104 to 85 × 1010 CFU/g, whereas total Salmonella count was found to be ranged from 16 × 106 to 18.5 × 1011 CFU/g, 15 × 104 to 44 × 107 CFU/g, and 13.2 × 105 to 21 × 1010 CFU/g fecal sample before starting sulfonamide treatment, during treatment with sulfonamide immediately after completion of sulfonamide treatment, respectively. The in vitro antibiotic sensitivity test showed that all the E. coli and Salmonella spp. isolated from diarrheic calves (100%) contained multidrug-resistant (MDR) phenotypes. Escherichia coli isolates were found 100% resistant to amoxicillin (AMX), cefuroxime, cephalexin (CN), erythromycin (ERY), and tetracycline (TET); whereas 94.4%, 86.1%, and 77.8% isolates were resistant to doxycycline (DOX), moxifloxacin (MOF), and gentamycin (GEN), respectively. In case of Salmonella isolates, all were found 100% resistant to AMX, CN, and ERY; whereas 91.7% of resistance was observed for DOX, MOF, cefuroxime, GEN, and TET. Based on the molecular screening of the antibiotic resistance genes, tetA gene was present in 83.3% of the isolated E. coli and 75% of the isolated Salmonella strains, whereas 83.3% E. coli and 79.2% Salmonella isolates contained blaTEM gene. Conclusion: These findings suggest that MDR E. coli and Salmonella spp. might be responsible for calf scouring, which is challenging to treat with antibiotics or sulfonamide drugs alone. Therefore, it is important to check the antibiotic sensitivity pattern to select a suitable antibiotic for the treatment of calf scoring. A suitable antibiotic or combination of an antibiotic and sulfonamide could be effective against E. coli and Salmonella spp. responsible for calf scouring.

Bacterial loads and antibiotic resistance profile of bacteria isolated from the most popular street food (Phuchka) in Bangladesh.

OBJECTIVE: Phuchka is one of the most common street foods in Bangladesh. It is served with salad, sweet and sour tamarind dispersed water, and minced eggs as topping at places where people usually gather. This makes these foods susceptible to bacterial contamination. Therefore, assessing the bacterial load and antimicrobial profile of organisms isolated from phuchka and other foodstuffs served with it was the focus of this study. MATERIALS AND METHODS: Bacterial isolates were isolated and identified from the samples after the bacterial loads were assessed as total viable count (TVC), total coliform count (TCC), and total staphylococcal count (TSC). The antibiotic resistance profile of the isolates was obtained using the disk diffusion method. Molecular detection of Escherichia coli isolates and the presence of gene responsible for tetracycline resistance was confirmed by polymerase chain reaction. RESULTS: According to the recommendations, the TVC value of 70% phuchka and egg samples was safe, whereas TSC value illustrated that 80% of both phuchka and egg samples were at safety level. For the TCC value, 80% egg and 70% phuchka samples were found to be safe for consumption. Among all the samples, the microbial loads of the vendors' hand wash were least in the safety level. Antibiotic sensitivity tests revealed that both Staphylococcus spp. and E. coli isolates were sensitive to gentamicin and ciprofloxacin but showed resistance to ampicillin. CONCLUSION: The data of this study indicate that phuchka can pose a public health problem as foodborne bacterial isolates which are antibiotic-resistant are found in it.

Isolation and Characterization of Multidrug-Resistant Escherichia coli and Salmonella spp. from Healthy and Diseased Turkeys.

Diseases caused by Escherichia coli (E. coli) and Salmonella spp. can negatively impact turkey farming. The aim of this study was to isolate and characterize multidrug-resistant (MDR) E. coli and Salmonella spp. in healthy and diseased turkeys. A total of 30 fecal samples from healthy turkeys and 25 intestinal samples from diseased turkeys that died of enteritis were collected. Bacterial isolation and identification were based on biochemical properties and polymerase chain reaction (PCR). Antibiogram profiles were determined by disk diffusion. The tetracycline-resistance gene tetA was detected by PCR. All samples were positive for E. coli. Only 11 samples (11/30; 36.67%) were positive for Salmonella spp. from healthy turkeys, whereas 16 (16/25; 64%) samples were positive for Salmonella spp. from diseased turkeys. E. coli isolated from diseased turkeys showed higher resistance to levofloxacin, gentamicin, chloramphenicol, ciprofloxacin, streptomycin, and tetracycline. Salmonella spp. isolated from healthy turkeys exhibited higher resistance to gentamicin, chloramphenicol, ciprofloxacin, streptomycin, imipenem, and meropenem. All E. coli and Salmonella spp. from both healthy and diseased turkeys were resistant to erythromycin. Salmonella spp. from both healthy and diseased turkeys were resistant to tetracycline. Multidrug resistance was observed in both E. coli and Salmonella spp. from diseased turkeys. Finally, the tetA gene was detected in 93.1% of the E. coli isolates and in 92.59% of the Salmonella spp. isolates. To the best of our knowledge, this is the first study to isolate and characterize tetA-gene-containing MDR E. coli and Salmonella spp. from healthy and diseased turkeys in Bangladesh. Both microorganisms are of zoonotic significance and represent a significant public health challenge.

Differential efficacies of marigold leaves and turmeric paste on the healing of the incised wound in sheep.

OBJECTIVE: This study was undertaken to compare the curative efficacy of marigold leaf paste and turmeric paste on healing the incised wound in sheep. The study also determined the antimicrobial effects and histopathological changes in a wound's healing process treated with these medicinal herbs. MATERIALS AND METHODS: Surgical wounds (n = 18) were created aseptically in the skin of the flank region of six healthy sheep dividing them into three experimental groups. Follow-up data were taken up to day 21. Different morphological characteristics of the wound and wound contraction (length and width) were recorded weekly. Samples were collected on days 1, 2, and 3 to test the antimicrobial effects and on days 1, 3, and 7 for histopathological studies. RESULTS: Treatment with marigold leaf paste and turmeric paste resulted in a swelled wound area of 11.78 ± 0.38 mm and 11.52 ± 0.27 mm, respectively. The wound areas were comparatively lower than that of the control group (11.44 ± 0.20 mm). Moreover, the least elevation (2.44 ± 0.12 mm) of the sutured line from the skin surface was noted and compared between the marigold leaf paste and normal saline (2.74 ± 0.13 mm). Bacterial colonies in the nutrient agar medium cultured with swabs from the normal saline-treated group's wound area were found on day 1. On the contrary, bacterial colonies were absent on days 2 and 3 of treatment in the groups treated with both the paste of marigold leaves and turmeric. Less intense tissue reactions and higher keratinization of epithelium were shown in the group treated with marigold leaf paste than turmeric paste and saline-treated groups. CONCLUSION: Marigold leaf paste showed less tissue reaction and healed the wounds effectively. Thus, this paste could be used for the treatment of superficial wounds in sheep.

Molecular detection of Vibrio cholerae from human stool collected from SK Hospital, Mymensingh, and their antibiogram.

OBJECTIVE: Vibrio spp., particularly, Vibrio cholerae is a major etiology of diarrhea in humans worldwide. In this study, we isolated and identified V. cholerae from the human stool of suspected cases along with antibiogram. MATERIALS AND METHODS: In total, 25 stool samples from cholera suspected patients were analyzed. Isolation and molecular detection of Vibrio species were performed based on staining, motility, cultural and biochemical characteristics followed by polymerase chain reaction (PCR) using groEL gene-specific primers. RESULTS: Among the 25 samples, seven showed growth of yellow color colonies on Thiosulfate-Citrate-Bile salts-Sucrose agar plates. The isolates were Gram-negative, curved shaped, and motile. Biochemically, they were found positive for indole and Methyl Red tests and negative for Voges-Proskauer test. Out of the seven positive samples, only three isolates were confirmed as Vibrio spp. using genus-specific primers. Subsequently, these three isolates were confirmed as V. cholerae by PCR using V. cholerae groEL gene-specific primers. Antibiotic sensitivity test revealed these three isolates as highly sensitive to azithromycin, chloramphenicol, gentamicin, and norfloxacillin while resistant to streptomycin, tetracycline, and oxacillin. CONCLUSION: Vibrio cholerae were isolated from the stool of diarrheic human patients and confirmed by PCR targeting the groEL gene. The isolates were found resistant to streptomycin, tetracycline and oxacillin, and need further characterization to reveal the molecular basis of their origin and resistance.

Beneficial effects of probiotic and phytobiotic as growth promoter alternative to antibiotic for safe broiler production.

OBJECTIVES: Drug residues in poultry products could lead to the development of antibiotic-resistant bacteria as in any living animal and human alike. Extensive use of antibiotics in animals to promote growth rate, increase feed efficiency, and for prevention of intestinal infections has led to the development of resistant bacteria in the gastrointestinal tract. The present study was conducted to evaluate the effects of biological supplementation of probiotic, phytobiotic, and their combination over antibiotic on growth performance, microbial load, and hematological parameters in Broiler. MATERIALS AND METHODS: Sixty-five broiler chicken were divided into five groups (12 birds in each group), namely, group A (basal diet), group B (antibiotic, Renamycin 100®), group C (phytobiotic, Galibiotic®), group D (probiotic, Bio-Top®), and group E (combination, Galibiotic®+Galibiotic®) and five were sacrificed for baseline data on day 0. RESULTS: Average final live weight gain was highest in group D (probiotic) than other groups. The feed conversion ratio was highest in group A and lowest in the probiotic group (group C). Blood samples were collected on 14th and 28th day for hematological studies. The mean hematology values regarding the total erythrocyte count, hemoglobin concentration, packed cell volume, and erythrocyte sedimentation rate differed significantly (p < 0.05) among groups. The pH of all the treatment groups was significantly decreased compared to the control group (p < 0.05) where group C was significantly (p < 0.05) lower than all other groups. Highest total viable cell count was observed in control (group A) and total coliform count in phytobiotic (group C) was significantly lower than in other treatment groups (p < 0.05). CONCLUSION: It may conclude that biological supplements have a significant positive impact on growth performance, hematological parameters, and gut microbial load in broiler chicken of which the probiotic showing the best effects. Supplementation of probiotic in feed could be one of the best candidates as an alternative to antibiotics as growth promoter for safe broiler production.

Dose-dependent response to phytobiotic supplementation in feed on growth, hematology, intestinal pH, and gut bacterial load in broiler chicken.

OBJECTIVE: The present study was aimed to evaluate dose-dependent effects of phytobiotic (Galibiotic) supplements in feed on growth performance, hematological parameters, intestinal pH, and gut bacterial population in broiler chick. MATERIALS AND METHODS: A total of 50 ten day old broiler chicks were divided into five groups, namely, Group A as control (without galibiotic), Group B (galibiotic at 1 gm/kg feed), Group C (galibiotic at 2 gm/kg feed), Group D (galibiotic at 5 gm/kg feed), and Group E (galibiotic at 10 gm/kg feed). All the birds were reared for 42 days and samples were collected before and after sacrifice. RESULTS: Live body weights showed no significant differences between the groups but overall feed conversion ratios (FCRs) of treatment groups were significantly low in Group E having the lowest. Blood samples collected for hematology differed significantly (p < 0.01) among the different groups. Intestinal pH was lower in treatment groups with Group E having the lowest. Cecal total viable count was highest in Group A and lowest in Group E. The cecal coliform count was low in all the treatment groups. CONCLUSION: Along with previously published report, it may conclude that the phytobiotic could be used as an alternative to antibiotics due to positive growth performance, lower FCR, carcass quality, and improved gut health of broiler chicks.

PCR-restriction fragment length polymorphism analysis using groEL gene to differentiate pathogenic Vibrio species.

Important pathogenic Vibrio species were differentiated by PCR-restriction fragment length polymorphism analysis. A 1117-bp groEL gene product was amplified using universal primers and digested using the restriction enzymes NruI or XbaI, revealing unique digestion patterns for each of the 10 Vibrio species, of which 7 were pathogenic in humans, along with 2 other species pathogenic in fish.

Multiplex PCR for the detection and differentiation of Vibrio parahaemolyticus strains using the groEL, tdh and trh genes.

Vibrio parahaemolyticus is a significant cause of human gastrointestinal disorders worldwide, transmitted primarily by ingestion of raw or undercooked contaminated seafood. In this study, a multiplex PCR assay for the detection and differentiation of V. parahaemolyticus strains was developed using primer sets for a species-specific marker, groEL, and two virulence markers, tdh and trh. Multiplex PCR conditions were standardised, and extracted genomic DNA of 70 V. parahaemolyticus strains was used for identification. The sensitivity and efficacy of this method were validated using artificially inoculated shellfish and seawater. The expected sizes of amplicons were 510 bp, 382 bp, and 171 bp for groEL, tdh and trh, respectively. PCR products were sufficiently different in size, and the detection limits of the multiplex PCR for groEL, tdh and trh were each 200 pg DNA. Specific detection and differentiation of virulent from non-virulent strains in shellfish homogenates and seawater was also possible after artificial inoculation with various V. parahaemolyticus strains. This newly developed multiplex PCR is a rapid assay for detection and differentiation of pathogenic V. parahaemolyticus strains, and could be used to prevent disease outbreaks and protect public health by helping the seafood industry maintain a safe shellfish supply.

Effect of a probiotic strain, Enterococcus faecium, on the immune responses of olive flounder (Paralichthys olivaceus).

The present study was aimed to investigate the effect of a probiotic, Enterococcus faecium, on the immune responses against infection with the marine fish pathogen Lactococcus garvieae in olive flounder (Paralichthys olivaceus). The immune responses were assessed by lysozyme activity, complement activity, protease activity, and expression of proinflammatory cytokines by RT-PCR. The lysozyme and complement activities were increased between 9 to 15 and 9 to 13 days, respectively, and antiprotease activity was slightly elevated after 5 days of probiotic treatment. The TNF-alpha and IL-1beta expressions were observed from kidney and spleen. The results of this study reveal that E. faecium induces immune-responsible materials and protects olive flounder from lactococcosis.