<div style="text-align:justify">Background: Antibiotic resistance is one of the major problems encountered in the therapy of canine pyometra. The ability of bacteria to form biofilm is implicated as one of the factors responsible for this. Escherichia coli and Staphylococcus aureus are the predominant bacteria associated with pyometra in canines and are known for their biofilm formation. Keeping in this view, a preliminary study was conducted to detect the biofilm forming strains of E. coli and S. aureus, if any, associated with canine pyometra. 
 
Methods: A total of 25 samples were collected, which included uterine discharges from cases of closed pyometra and anterior vaginal swabs from open pyometra. The isolates of E. coli and S. aureus were identified based on the cultural, morphological and biochemical characteristics. These isolates were subjected to antibiotic sensitivity test employing disc diffusion method. For biofilm detection, the isolates were screened by Congo red agar method, tube method and tissue culture plate method. 
 
Result: From 25 samples, two Streptococcus spp., thirteen&nbsp;Staphylococcus spp., seven E.coli, five Klebsiella spp. and two Pseudomonas spp. were isolated. All the isolates were found to be multi-drug resistant on antibiogram. The tissue culture plate and Congo red agar method was found more sensitive to detect the biofilm formation by S. aureus and E.coli isolates, respectively. The biofilm forming strains showed higher degree of antibiotic resistance in comparison with non-formers, indicating it as one of the major reasons for failure of antibiotic therapy in canine pyometra.</div>

Detection of Biofilm Formation by Escherichia coli and Staphylococcus aureus Associated with Canine Pyometra

<div style="text-align: justify;">The aim of the study was to detect Salmonella from coriander leaves by polymerase chain reaction, determine biofilm formation and antimicrobial resistance profiling of the isolates. Four Salmonella isolates were recovered from the total of 60 samples. The isolates were confirmed by PCR. The confirmed isolates were subjected for biofilm production assay and showed that all Salmonella isolates were able to form biofilm. Biofilm forming isolates were also subjected for antimicrobial resistance profiling. Three isolates were showed sensitivity against norfloxacin (92.5%). One was showing sensitivity against cefotaxime (90%). All the four isolates were showing complete resistant to ciprofloxacin, nalidixicacid, chloramphenical, gentamicin and cephalexin. Clindamycin and ofloxacin were showed sensitivity 20%. The finding of this study shows that coriander leaves are potential host for the transmission of Salmonella to human and animals. The ability of the isolates to form biofilm reveals the potential of the isolates to persist on the green leafy vegetables and the pathogenic status of the isolates as well as ability to resist antimicrobial chemotherapy.</div>

PCR based detection and Biofilm formation of Salmonella from fresh coriander leaves (Coriandrum sativum)

<div>Background:&nbsp;Escherichia coli and Klebsiella pneumoniae are two important commensals with pathogenic potentiality in animals and birds, causing intestinal and extra-intestinal infections. These are somewhat zoonotic too via different routes. So, the present study was undertaken to investigate the prevalence of these two bacteria from cloaca and environment of ducks from different districts of West Bengal, followed by detailed characterization and antibiogram. 
 
Methods: Around 197 duck and environmental samples were collected from different districts of West Bengal. E. coli and K. pneumoniae were isolated from collected samples as per standard methods followed by serotyping of the E. coli isolates. The detection of ESBL/ACBL producing isolates was done both phenotypically and genotypically by detecting the presence of antibiotic-resistant genes. Virulence property and biofilm-producing ability of these isolates were also studied. Antibiogram of the ESBL and biofilm positive isolates were done using common 12 antibiotics by disc diffusion method.&nbsp; &nbsp; 
 
Result: A total of 134(68.02%) Escherichia coli and 33(16.75%) Klebsiella pneumoniae isolates were recovered and confirmed by PCR. The prevalence of E. coli and K. pneumoniae in cloacal swabs were less in comparison to the environmental samples (72.09% and 20.93%). Approx. 79.85% E. coli and 72.73% K. pneumoniae isolates were found to be the ESBL/ACBL producers in vitro. E. coli and K. pneumoniae isolates possessed all major antibiotic-resistant genes with the blaAmpC (64.18% &amp; 48.48%) being the most prevalent one. Twenty three (17.16%) E. coli isolates were found positive for the virulence genes, of which stx1 (39.13%) was the most common one as detected by multiplex PCR. No virulence gene was found in K. pneumoniae isolates. Ninety one (67.91%) E. coli and 26(78.79%) K. pneumoniae isolates possessed the biofilm-producing genes. All the ESBL and biofilms producing E. coli and K. pneumoniae isolates showed 100% - 95.24% resistance to ampicillin. A high percentage of resistances were also seen against cefotaxime, ticarcillin/ clavulanic acid, ertapenem, ceftazidime, norfloxacin and ciprofloxacin, which may be due to several reasons. Maximum overall sensitivity was detected against chloramphenicol, co-trimoxazole and gentamicin.</div>

Detection and Characterization of &beta;​-lactamases and Biofilm-producing Escherichia coli and Klebsiella pneumoniae from Ducks and Duck Environments in West Bengal

<div>Salmonella has emerged as one of the most important food-borne pathogens for humans as well as animals and the ability of biofilm formation by these bacteria has further aided their survival in unfavorable environment. Characterization of these biofilm producing bacteria isolated from pigs and chicken may lead to formulation of strategies for prevention and control of Salmonella infections. Therefore, the present study was conducted to isolate Salmonella from pigs and poultry of Mizoram, determine their biofilm producing ability by phenotypic and genotypic methods along with their virulence and antimicrobial resistance properties. A total of 15 Salmonella spp. (pig=9, poultry=6) was isolated from 100 faecal samples from pigs and 50 cloacal swabs from poultry and biofilm producing ability of the isolates was determined by microtiter plate assay. A total of 10 (66.67%) isolates were found to be biofilm producer. All the biofilm producing bacterial isolates were investigated for antimicrobial sensitivity and distribution of selected biofilm associated genes (csgA, csgD and adrA), virulence genes (invA, stn and sefA) and antimicrobial resistance (AMR) genes (blaTEM, blaSHV and blaCTX-M). The most prevalent resistance was found against ceftazidime (80%), ceftriaxone (80%), cefixime (70%), cefotaxime (70%), gentamicin (70%), cotrimoxazole (60%) and ampicillin (60%). A total of 7 (70%) isolates were resistant to at least three different classes of antimicrobial agents and considered as multidrug resistant. All the isolates were positive for adrA (100%) but negative for csgA and csgD genes. The most frequent virulence gene was invA (100%) and stn (100%). Among the AMR genes, blaTEM &nbsp;(60%) was found to be the major AMR determinants. Moreover, a total of 7 Salmonella isolates were &nbsp; positive for at least one of t biofilm associated genes, virulence genes and AMR genes.&nbsp;</div>

<div>Molecular detection of biofilm, virulence and antimicrobial resistance associated genes of Salmonella serovars isolated from pig and chicken of Mizoram, India</div>

<div>Background: Bovine mastitis is a multifactorial and is one of the most challenging disease. It can be caused by many different bacterial species, the most common of which are Staphylococcus species and Streptococcus species. The prevalence of different species varies temporally, geographically and also due to control measures adopted in herds. The present study is on the biofilm forming Streptococcus uberis causing mastitis and the effect of antibiofilm agents on the antimicrobial resistance of the microorganisms.&nbsp; 
 
Methods: The isolates were identified by Polymerase chain reaction (PCR). Two antibiofilm agents, resveratrol and ursolic acid (UA) each at two concentrations (30 &mu;g/ml, 100 &mu;g/ml) were used for the study. Biofilm formation and rate of biofilm inhibition was detected using quantitative microtiter plate (MTP) assay and biofilm gene (lux S) was detected using PCR. The isolates treated with antibiofilm agents were subjected to standard disc diffusion test with 7 antibiotics and the change in antibiotic resistance was studied.&nbsp; 
 
Result: The mean&plusmn;SE values of inhibition rates of 29 S. uberis isolates by 30 &mu;g/ml UA, 100 &mu;g/ml UA, 30 &mu;g/ml resveratrol, 100 &mu;g/ml resveratrol were 33.96&plusmn;3.17%, 57.40&plusmn;2.8%, 31.35&plusmn;3.12% and 46.28&plusmn;3.47%, respectively. Biofilm inhibiting agents along with all antibiotics had reduced antimicrobial resistance by 1.5-2 times on in vitro antibiotic resistance testing by disc diffusion at 100 &mu;g/ml concentration. The antibiofilm agents were found to be very effective to control antibiotic resistance of S. uberis from mastitic milk samples in vitro.</div>

​Efficacy of Resveratrol and Ursolic Acid on Biofilm Inhibition and Antimicrobial Resistance of Streptococcus uberis

<div>Background: Methicillin resistant Staphylococcus aureus (MRSA) refers to a group of gram-positive bacteria that are genetically distinct from other strains of Staphylococcus aureus that has developed, through horizontal gene transfer, natural selection and multiple drug resistance to beta lactam antibiotics and thus causing many severe diseases which are very problematic to treat as they are resistant to different antibiotics. The above situation is further aggravated by the formation of biofilms which are structured aggregation of surface attached microbes encased in an extracellular matrix. Thus, the time required to develop biofilms becomes an important point to study.&nbsp; 
 
Methods: In the present study, detection of the extent of biofilm formation by MRSA isolates, was performed using the Microtitre Plate Assay. The test was performed in triplicates and was continued for a time period of 7 days to detect the extent of biofilm formation with increase in incubation interval. 
 
Result: On comparing the biofilm forming ability of different isolates from Day 1 to Day 7, it could be clearly observed that the biofilm forming capacity of isolates gradually increased with increase in incubation time and most of the isolates became strong biofilm producers from Day 4.</div>

Effect of Incubation Time on the Biofilm Formation by Methicillin Resistant Staphylococcus aureus

<div>Background: Streptococcus uberis (S. uberis) is an environmental pathogen causing mastitis in Dairy cattle. It causes recurrent mastitis and reduction in milk production in livestock causing economic loss. The prevalence of S. uberis intramammary infections is due to ability of the organism to form biofilm in udder tissue. The present study is on in-vitro biofilm production, the correlation of luxS gene and the biofilm formation in S. uberis.&nbsp; 
 
Methods: A total of 91 mastitic milk samples were collected from cattle and buffaloes brought at Veterinary Hospitals and farms in Krishna, Guntur and West Godavari districts, Andhra Pradesh. The identification of the culture isolates was based on cultural and biochemical characteristics and confirmed by Polymerase Chain Reaction (PCR). The Streptococcus species cultures showing greyish, pinpointed colonies and/or aesculin hydrolysis on Edwards medium were further identified by various biochemical tests viz., catalase test, ninhydrin test, sodium hippurate hydrolysis test and type of haemolysis on 7% sheep blood agar. Confirmation of the isolates by PCR was followed by detection of biofilm formation using qualitative Congo red agar (CRA) method, quantitative microtiter plate (MTP) assay and biofilm gene (luxS) was detected using PCR. 
 
Conclusion: From this study it is suggestable that for biofilm study both phenotypic and genotypic methods should be taken together which can be influenced by various other factors also. MTP assay was a good choice for quantitative biofilm determination, which was giving a more accurate and understandable results. The results express that any of the isolates without luxS didn&rsquo;t produce a strong biofilm and it is concluded that there may be other genes for regulation of biofilm production and/or luxS gene has a regulatory role for one or more genes related to biofilm formation in S. uberis.</div>

<div>luxS Gene and Biofilm Formation in Streptococcus uberis Isolated from Bovine Mastitis Cases</div>

<div style="text-align: justify;">Pseudomonas aeruginosa is considered as the most potent member of multidrug-resistant (MDR) and extremely resistant (XDR) gram-negative pathogens (&lsquo;ESCAPE&rsquo; group) emerged throughout the world with the property to &lsquo;escape&rsquo; the treatment with antibiotics. Biofilm formation is a significant virulence property of P. aeruginosa generating not only antibiotic resistance, but also it acts as a constant source of infection in the host and it can prevent host defence such as chemotaxis of polynuclear immune cells. There is paucity of scientific literatures regarding characterization of biofilm producing P. aeruginosa isolated from livestock and birds. A total of 200 rectal swabs were collected from pigs (n=100) and chickens (n=100) from Mizoram state of India. All the specimens were processed of isolation and identification of P. aeruginosa, which were further confirmed by 16S rRNA PCR and Phoenix bacterial identification system. All the isolates were subjected to detection of biofilm producing ability by microtiter plate assay, antimicrobial sensitivity by disc diffusion assay and detection of selected biofilm producing genes as well as selected beta lactamase genes by specific PCR assay. A total of 11 P. aeruginosa were isolated from pigs (n=6) and chickens (n=5). All the isolates were recorded as positive for biofilm production by microtiter plate assay and were positive for at least one biofilm associated genes. A total of 8 isolates were positive for blaTEM gene and one isolates was positive for blaCTX-M gene. This is probably the unique report on isolation of P. aeruginosa from animals carrying both beta lactamase as well as biofilm producing genes.</div>

<div style="text-align: justify;">Molecular Characterization of Biofilm-Producing Pseudomonas aeruginosa Isolated from Healthy Pigs and Chicken in India</div>

<div>Background: The present study assessed the occurrence and biofilm forming potential of ESBL-producing Escherichia coli isolates recovered from raw salad vegetables collected from 32 retail vegetable shops of Mirzapur district, Uttar Pradesh, India (n=224). 
&nbsp; 
Methods: Standard bacteriological culture methods using cefotaxime-supplemented EMB agar were used for initial isolation of cefotaxime-resistant E. coli which were subjected to phenotypic detection of ESBL production by Kirby-Bauer disc diffusion test. Subsequently, all the phenotypically positive ESBL-producing E. coli isolates were subjected to determine their biofilm-producing ability by Congo red agar (CRA) assay and microtiter plate assay (MPA) followed by genotypic confirmation of biofilm production by PCR assay targeted at fim-H, Sfa and papC genes. Additionally, the isolates were genotypically explored for a potential presence of Shiga-like toxin genes (stx1 and stx2). 
&nbsp; 
Result: Altogether, 276 cefotaxime-resistant E. coli isolates were recovered, of which, 99.27% (274/276) were observed to be phenotypically positive for ESBL production. The brown, dry and rough (BDAR) and red, dry and rough (RDAR) colony morphotypes were observed by 70.80% (194/274) and 29.19% (80/274) of the isolates, respectively by CRA method revealing their potential to form biofilms. Biofilm formation was evident in 94.16% (258/274) of the isolates by MPA method, of which 79.45% (205/258), 17.05% (44/258) and 3.48% (9/258) were considered as weak, moderate and strong biofilm producers, respectively.&nbsp; The fimH gene was found to be the predominant genetic determinant of biofilm formation which was detected in 77% (211/274) of the tested isolates followed by Sfa gene (11.31%) and papC gene (4.01%). A significant difference (p&lt;0.05) was evident between CRA and MPA methods as well as phenotypic and genotypic methods for detection of biofilm formation among the recovered isolates. In addition, significant differences (p&lt;0.05) were also observed upon vegetable-wise comparison of ESBL-producing E. coli isolates and the presence of biofilm producing genes. The stx1 and stx2 genes were found to be present in one isolate recovered from a carrot sample and two isolates from coriander samples, respectively. . The findings of this study highlight the widespread occurrence of ESBL-producing E. coli from raw salad vegetables with significant biofilm-forming potential posing a public health risk. The detection of Shiga-like toxin genes in some isolates further underscores the need for stringent surveillance and improved hygiene practices to mitigate contamination risk in fresh produce.</div>

Assessment of Biofilm Formation and Shiga-like Toxin Genes in ESBL-producing E. coli Isolates Recovered from Retail Salad Vegetables in Mirzapur District, Uttar Pradesh, India

<div>Background: Streptococcus suis is a zoonotic pathogen closely related to the respiratory tract of swine. Paeonol is a traditional Chinese medicine with antibacterial effect. This study demonstrated the effect of paeonol on biofilm formation and the expression of virulence factors of Streptococcus suis.&nbsp; 
 
Methods: The minimum inhibitory concentration (MIC) of paeonol on Streptococcus suis was determined by double dilution method. Crystal violet staining method was used to determine the effect of paeonol on the biofilm formation. The effects of paeonol on the morphology and structure of Streptococcus suis biofilm were observed by field emission scanning electron microscopy (FE-SEM). Real-time quantitative PCR (qRT-PCR) was used to detect the expression levels of Streptococcus suis related virulence factors under sub-MIC (1/4 MIC, 1/8 MIC) paeonol. 
 
Result: The results showed the effect of 1/4 MIC and 1/8 MIC paeonol on biofilm formation and mature biofilm was significant (*P&lt;0.05). Under the action of paeonol, the biofilm structure becomes sparse. qRT-PCR results showed that 1/8 MIC paeonol inhibits bacterial adhesion by up-regulating the expression of mrp virulence genes, thereby inhibiting the formation of Streptococcus suis biofilm.</div>

Detection of Biofilm Formation by Escherichia coli and Staphylococcus aureus Associated with Canine Pyometra

PCR based detection and Biofilm formation of Salmonella from fresh coriander leaves (Coriandrum sativum)

Detection and Characterization of β-lactamases and Biofilm-producing Escherichia coli and Klebsiella pneumoniae from Ducks and Duck Environments in West Bengal

Molecular detection of biofilm, virulence and antimicrobial resistance associated genes of Salmonella serovars isolated from pig and chicken of Mizoram, India

Efficacy of Resveratrol and Ursolic Acid on Biofilm Inhibition and Antimicrobial Resistance of Streptococcus uberis

Effect of Incubation Time on the Biofilm Formation by Methicillin Resistant Staphylococcus aureus

luxS Gene and Biofilm Formation in Streptococcus uberis Isolated from Bovine Mastitis Cases

Molecular Characterization of Biofilm-Producing Pseudomonas aeruginosa Isolated from Healthy Pigs and Chicken in India

Assessment of Biofilm Formation and Shiga-like Toxin Genes in ESBL-producing E. coli Isolates Recovered from Retail Salad Vegetables in Mirzapur District, Uttar Pradesh, India

Effects of Paeonol Sub-inhibitory Concentration on Streptococcus suis Biofilm and Expression of Virulence Genes