Histochemical characterization of duodenum at early, mid and late prenatal period in goats

Goats [Capra aegagrus hircus (L.)] play a significant role in providing supplementary income and livelihood to humans. The intestine plays a major role in foetus development and growth, and duodenum, as the part of small intestine, is responsible for breakdown of food. As there is not much studies available in literature on this aspect, here, we investigated the developing duodenum of 30 goat embryos/foeti irrespective of breed and sex. The tissues were fixed in 10% neutral buffered formalin and in cold acetone. Different histochemical techniques were applied for the detection and localization of neutral mucopolysaccharides, acid mucopolysaccharides, bound lipids, alkaline phosphatase enzymes, acid phosphatase enzymes, and DNA. The intensity of reactions increased as the age of foeti advanced. The goblet cells of the intestinal gland showed moderate reaction for Periodic acid schiff (PAS) and Acid mucopolysaccharides (AMPs) in the mid prenatal period (Gr. II) and intense to highly intense reaction in the late prenatal period (Gr. III). The cytoplasm of the epithelial cells exhibited negative to mild reaction for bound lipids in Gr. I, mild to moderate in Gr. II and mild to intense reaction in Gr. III. The luminal border of epithelial cells exhibited mild reaction in Gr. I & II, and moderate to intense reaction in Gr. III. The luminal border showed weak acid phosphatase reaction in Gr. I & II, and mild to moderate reaction in Gr. III. The nuclei of the epithelial cells showed mild to moderate Feulgen reaction in Gr. I & II, and intensely positive reaction in Gr. III.

Cytodifferentiation of fundic part of glandular stomach in non descript breed of Indian prenatal goat

Though the anatomy and physiology of the adult caprine (Capra hircus L.) stomach have been investigated extensively, the early development of the abomasum has not yet been fully elucidated. The glandular part of abomasum plays an important role in digestion of ingested food by action of gastric juices. Very few studies have been conducted so far regarding histogenesis of goat foetal abomasum in India. In the present study, we have investigated the embryonic and early foetal development of the goat, Capra hircus L. fundic abomasum. We collected 36 developing abomasum of healthy and normal embryos/foeti of goat and assigned them into three group viz. Gr. I (0-50 days of gestation), Gr. II (51-100 days of gestation) and Gr. III (101-150 days of gestation). Small pieces of tissues were processed by routine paraffin. The wall of glandular stomach, the fundic part, was composed of epithelium, pleuripotent blastemic tissue and serosa up to 44 days of gestation. Tunica muscularis became separable at 46 days of gestation. The epithelium was stratified type up to 50 days and gradually changed to pseudo-stratified columnar to simple columnar type from 76 days of gestation. Primary and secondary abomasal folds were observed at 51 and 76 days of gestation, respectively. Gastric pit, the fore runner of gastric gland was reported first at 70 days. The gland became branched tubular type at 145 days. The cells found in the mucosa of the abomasum were surface epithelial cells, chief cells, parietal cells, mucous neck cells and undifferentiated cells. Chief and parietal cell were observed at 76 days and mucous neck cells at 82 days of gestation. Reticular, collagen and elastic fibers came into sight at 38, 76 and 100 days of gestation, respectively. The present study is expected to supplement known data and knowledge regarding histogenesis of goat fetal abomasum and help in diagnosis and treatment of related congenital anomalies.

Methicillin resistant Staphylococcus aureus (MRSA) in India Prevalence & susceptibility pattern.

Background & objectives: Methicillin resistant Staphylococcus aureus (MRSA) is endemic in India and is a dangerous pathogen for hospital acquired infections. This study was conducted in 15 Indian tertiary care centres during a two year period from January 2008 to December 2009 to determine the prevalence of MRSA and susceptibility pattern of S. aureus isolates in India. Methods: All S. aureus isolates obtained during the study period in the participating centres were included in the study. Each centre compiled their data in a predefined template which included data of the antimicrobial susceptibility pattern, location of the patient and specimen type. The data in the submitted templates were collated and analysed. Results: A total of 26310 isolates were included in the study. The overall prevalence of methicillin resistance during the study period was 41 per cent. Isolation rates for MRSA from outpatients, ward inpatients and ICU were 28, 42 and 43 per cent, respectively in 2008 and 27, 49 and 47 per cent, respectively in 2009. The majority of S. aureus isolates was obtained from patients with skin and soft tissue infections followed by those suffering from blood stream infections and respiratory infections. Susceptibility to ciprofloxacin was low in both MSSA (53%) and MRSA (21%). MSSA isolates showed a higher susceptibility to gentamicin, co-trimoxazole, erythromycin and clindamycin as compared to MRSA isolates. No isolate was found resistant to vancomycin or linezolid. Interpretation & conclusions: The study showed a high level of MRSA in our country. There is a need to study epidemiology of such infections. Robust antimicrobial stewardship and strengthened infection control measures are required to prevent spread and reduce emergence of resistance.

AmpC β-lactamases in nosocomial isolates of Klebsiella pneumoniae from India.

Background & objectives: AmpC β-lactamases are clinically significant since these confer resistance to cephalosporins in the oxyimino group, 7-α methoxycephalosporins and are not affected by available β-lactamase inhibitors. In this study we looked for both extended spectrum β-lactamases (ESBL) and AmpC β-lactamases in Klebsiella pneumoniae clinical isolates. Methods: One hundred consecutive, non-duplicate clinical isolates of K. pneumoniae collected over a period of one year (June 2008 - June 2009) were included in the study. An antibiotic susceptibility method was used with 10 antibiotics for Gram-negative infections which helped in screening for ESBL and AmpC β-lactamases and also in confirmation of ESBL production. The detection of AmpC β-lactamases was done based on screening and confirmatory tests. For screening, disc diffusion zones of cefoxitin <18 mm was taken as cefoxitin resistant. All cefoxitin resistant isolates were tested further by AmpC disk test and modified three dimensional test. Multiplex-PCR was performed for screening the presence of plasmid-mediated AmpC genes. Results: Of the 100 isolates of K. pneumoniae studied, 48 were resistant to cefoxitin on screening. AmpC disk test was positive in 32 (32%) isolates. This was also confirmed with modified three dimensional test. Indentation indicating strong AmpC producer was observed in 25 isolates whereas little distortion (weak AmpC) was observed in 7 isolates. ESBL detection was confirmed by a modification of double disk synergy test in 56 isolates. Cefepime was the best cephalosporin in synergy with tazobactam for detecting ESBL production in isolates co-producing AmpC β-lactamases. The subsets of isolates phenotypically AmpC β-lactamase positive were subjected to amplification of six different families of AmpC gene using multiplex PCR. The sequence analysis revealed 12 CMY-2 and eight DHA-1 types. Interpretation & conclusions: Tazobactam was the best β-lactamase inhibitor for detecting ESBL in presence of AmpC β-lactamase as this is a very poor inducer of AmpC gene. Amongst cephalosporins, cefepime was the best cephalosporin in detecting ESBL in presence of AmpC β-lactamase as it is least hydrolyzed by AmpC enzymes. Cefepime-tazobactam combination disk test would be a simple and best method in detection of ESBLs in Enterobacteriaceae co-producing AmpC β-lactamase in the routine diagnostic microbiology laboratories.

Phenotypic method for differentiation of carbapenemases in Enterobacteriaceae: Study from north India.

Aims: Carbapenems are usually the choice of antimicrobials in infections caused by Enterobacteriaceae bacteria-producing ESBL (extended spectrum β-lactamases) and Amp C. Resistance to carbapenems is mostly due to production of enzymes - Carbapenemases, which are divided into Ambler Classes A, B and D. Phenotypic detection and differentiation of types of Carbapenemases in carbapenem-resistant Enterobacteriaceae (CRE) is important for proper infection control and appropriate patient management. Materials and Methods: The present study done in a tertiary care hospital from North India differentiates Class A (KPC type) and B (MBL type) carbapenemases among Enterobacteriaceae isolates by simple phenotypic method that uses both the inhibitors EDTA and phenylboronic acid. Results: Total of 330 strains of Enterobacteriaceae were included in the study. Out of these 330 strains, 26 strains were resistant to carbapenems. The prevalence of CRE in our Institute is 7.87% (26/330). Conclusions: The prevalence of Enterobacteriaceae strains producing MBL type carbapenemase in our health care setup is 5.75% (19/330). None of the strains among the carbapenem-resistant bacterial isolates showed production of KPC enzyme. The need of the hour is simple, rapid and cost effective tests which will be able to identify and distinguish resistant pathogens for improved patient outcome, facilitating efficient infection control and reducing the escalation of resistance.

Screening & profiling of quorum sensing signal molecules in Pseudomonas aeruginosa isolates from catheterized urinary tract infection patients.

Background & objectives: Catheter associated urinary tract infections are the second most common nosocomial infections and Pseudomonas aeruginosa is the third most common organism responsible for these infections. In this study P. aeruginosa isolates from catheterized urinary tract infection patients were screened and profiled for the presence of different type of quorum sensing (QS) signal molecules. Methods: Screening and quantitation of AHLs was done by using cross feeding assay and by determining β-galactosidase activity respectively using Escherichia coli MG4 as reporter strain. Further, AHL profiles were determined by separating AHLs on TLC coupled with their detection using Chromobacterium violaceum CV026 and Agrobacterium tumifaciens A136 biosensor strains. Results: All uroisolates from catheterized patients having urinary tract infections were found to be producers of QS signal molecules. There were differences in amounts and type of AHL produced amongst uroisolates of P. aeruginosa. Several AHLs belonging to C4-HSL, C6-HSL, oxo-C6-HSL, C8-HSL, C10-HSL and C12-HSL were determined in these strains. Interpretation & conclusions: Simultaneous use of more than one reporter strain and assay method proved useful in determining the AHLs profile in uroisolates of P. aeruginosa. Observed differences in the amounts and types of AHLs may reflect differences in virulence potential of P. aeruginosa to cause UTIs which can be further confirmed by employing animal model system. The present study speculates that production of QS signal molecules may act as a new virulence marker of P. aeruginosa responsible for causing catheter associated UTIs and can be considered as futuristic potential drug targets towards treatment of UTIs.

Epidemiology & antibiograms of Vibrio cholerae isolates from a tertiary care hospital in Chandigarh, north India.

Background & objectives: Cholera is endemic in Chandigarh and its surrounding areas. This retrospective study was undertaken over a period of nine years (January 1999-December 2007) from a tertiary care hospital in north India to understand the changing epidemiology aspects and antibiotic resistance patterns in Vibrio cholerae isolates. Methods: A total of 277 isolates of V. cholerae were included in the study. V. cholerae was identifi ed by standard microbiological procedures. Antibiotic sensitivity testing was performed by disc diffusion method and isolates phage typed. Results: All the isolates were identifi ed as V. cholerae O1 biotype El Tor serotype Ogawa; phage 27 was the predominant type. Men were more commonly affected with maximum number in the age group 0-5 yr. Majority of the isolates were resistant to furazolidone but sensitive to gentamicin and cefotaxime. Resistance pattern to amoxycillin was variable. Three isolates were found to be resistant to ciprofl oxacin. All the patients presented during June-October coinciding with the monsoon season and a majority were from suburbs. Interpretation & conclusions: The emergence of resistance amongst V. cholerae especially towards ciprofl oxacin may signifi cantly infl uence the control strategies in future outbreaks. Phage 27 remained the predominant type in all the years. Continuous surveillance with regard to drug resistance, early detection and a strong regional commitment may help contain the disease.

An increase in enteric fever cases due to Salmonella Paratyphi A in & around Chandigarh.

BACKGROUND & OBJECTIVE: Enteric fever is a major public health problem in India. It is classically caused by Salmonella enterica serotype Typhi. Salmonella enterica serotype Paratyphi A which had been reported less frequently from cases of enteric fever has shown an increasing trend since 1996 in India. There is also variation in the antimicrobial susceptibility of Salmonella Paratyphi A from different parts of the country. An attempt is therefore made to study the rate of isolation and antimicrobial susceptibility pattern of Salmonella Paratyphi A from cases of enteric fever coming to a tertiary care hospital at Chandigarh. METHODS: The blood samples of patients suspected of having enteric fever and admitted to Government Medical College and Hospital, Chandigarh, from January 2006 to April 2007 (11,240) were processed by conventional methods. Antimicrobial susceptibility was tested by Kirby-Bauer disc diffusion method. The minimum inhibitory concentration to two antibiotics- ciprofloxacin and chloramphenicol was determined by agar dilution technique. Simultaneously, retrospective analysis was done from January 2003-December 2005 to study any difference in the incidence and antimicrobial susceptibility pattern of Salmonella Paratyphi A among enteric fever patients. RESULTS: Of 305 total isolates, 231 were S. Typhi and 84 S. Paratyphi A rise. The number of Salmonella Paratyphi A cases rose from 27 in 2006 (34.18%) to 13 (40.63%) in four months of 2007. All were sensitive to ciprofloxacin and cefotaxime but MIC to ciprofloxacin was raised (0.125-0.5 microg/ml). Resistance to nalidixic acid was 92.5 per cent. Chloramphenicol sensitivity re-emerged with 90 per cent isolates sensitive to it while sensitivity to ampicillin dropped (72.5%) as compared to previous years. Only one isolate was multi-drug resistant. INTERPRETATION & CONCLUSION: The present study conferencing Salmonella Paratyphi A as the rapidly emerging pathogen of enteric fever. With increasing resistance to fluoroquinolones and possibility of re-emergence of sensitivity to chloramphenicol, the policy of empirical treatment of enteric fever needs to be rationalized.

Isolation of Salmonella paratyphi A from renal abscess.

Intrarenal abscesses remain a significant cause of morbidity and mortality as well as a diagnostic dilemma because a plethora of microorganisms can cause this condition. A definitive diagnosis is made by demonstrating the organisms from the aspirate and the success or failure of therapy depends upon the antimicrobial sensitivity pattern. Enteric fever is a multisystem disorder caused by invasive strains of salmonella. Salmonellosis continues to be a major public health problem, especially in developing countries. Classic enteric fever is caused by S. typhi and usually less severe enteric fevers are caused by S. paratyphi A, B, or C. However, at times S. paratyphi is capable of causing serious and often life-threatening infections like infective endocarditis, pericarditis, empyma, sino-venous thrombosis, osteomyelitis, meningitis, bone marrow infiltration, hepatitis and pancreatitis. There are anecdotal case reports in world literature of abscesses being caused by this organism. Renal involvement like bacteriuria, nephrotic syndrome and acute renal failure have been reported due to S. parayphi A. S. paratyphi A has never been implicated in renal abscess, we report one such case that was managed successfully with medical therapy.

Avian Influenza: Current Scenario.

Avian influenza commonly called as 'Bird flu' is a matter of great concern which has caused three major pandemics in twentieth century killing millions of people. Recent epidemic with H5N1 type strain started in 1997 and is still continuing. Disease has assumed new proportions since its confirmed association with migratory birds. In India also, three separate outbreaks have been announced till now by Government of India. This virus has tremendous capacity for gene reassortment and humans by enlarge do not have any immunity against them. This fact has raised global fears of an imminent influenza pandemic. The neuraminidase inhibitors currently are the only drugs which can be used, in case pandemic occurs. Therefore, timely development of an effective influenza vaccine is must and should be made a public health priority.

An update on newer beta-lactamases.

The resistance to beta-lactam antibiotics is an increasing problem worldwide and beta lactamases production is the most common mechanism of drug resistance. Both global and Indian figures showed a marked increase in the number of beta-lactamases producing organisms. These enzymes extended spectrum beta-lactamases (ESBLs) are numerous and continuous mutation has led to the development of enzymes having expanded substrate profile. To date, there are more than 130 TEM type and more than 50 sulphydryl variable (SHV) type beta-lactamases found in Gram negative bacilli. ESBL producing Enterobacteriaceae are, as a rule, resistant to all cephalosporins and extended spectrum penicillins including the monobactam, aztreonam, while resistance to trimethoprim - sulphamethaxazole and aminoglycosides is frequently co-transferred on the same plasmid. Many ESBL producing organisms also express Amp C beta-lactamases. Amp C- beta-lactamases are clinically significant, as these confer resistance to cephalosporins in the oxyimino group, 7 alpha-methoxy cephalosporins, and are poorly inhibited by clavulanic acid. Carbepenems are the drugs of choice for the treatment of infections caused by ESBL producing organisms but carbapenemases (MBLs) have emerged and have spread from Pseudomonas aeruginosa to Enterobacteriaceae. The routine clinical microbiology laboratories should employ simple methods to recognize these enzymes using various substrates and inhibitors. These organisms may lead to therapeutic dead ends. Presently, the therapy relies on beta-lactam/ beta-lactamases inhibitor combinations, carbepenems and piperacillin - tazobactam plus aminoglycoside combination. Proper infection control practices and barrier precautions are essential to contain the organisms producing beta-lactamases.

Comparative in vitro activities of seven new beta-lactams, alone and in combination with beta-lactamase inhibitors, against clinical isolates resistant to third generation cephalosporins

We examined the drug susceptibility pattern of Gram-negative bacilli to seven new beta-lactams. A total of 277 non-duplicate gramnegative bacilli strains belonging to the Enterobacteriaceae family, Pseudomonas and Acinetobacter species, isolated from various clinical samples were tested for susceptibility to imipenem, piperacillin/tazobactam, cefoperazone/sulbactam, ticarcillin/clavulanate, cefdinir, cefepime and cefpirome with the disk diffusion technique. The percentage resistance was low for imipenem (7.2 percent), piperacillin/tazobactam (2.8 percent), cefoperazone/sulbactam (5.4 percent). However, a high frequency of resistance was observed to ticarcillin/clavulanate (83.9 percent), cefdinir (70.6 percent), cefepime (45.5 percent) and cefpirome (84.4 percent). We conclude that imipenem, piperacillin/tazobactam and cefoperazone/sulbactam are effective antibiotics in our environment, whereas ticarcillin/clavulanate, cefdinir, cefepime and cefpirome are relatively uneffective.

Phenotypic characters of urinary isolates of Pseudomonas aeruginosa & their association with mouse renal colonization.

BACKGROUND & OBJECTIVE: Very few studies regarding production of virulence factors in different predominant serotypes of uropathogenic Pseudomonas aeruginosa are available and they have not been correlated to in vivo pathogenicity in the urinary tract. This study was carriedout with the objective to analyze the phenotypic characters of uroisolates of P. aeruginosa in vitro and to study the association of these virulence traits with their ability to cause nephropathogenicity in mouse model of ascending urinary tract infection (UTI). METHODS: Protease, elastase, alginate, haemolysin, pyochelin, pyoverdin and phopholipase C were measured using standard protocols in 18 uroisolates of P. aeruginosa isolated from patients suffering from complicated UTIs. An ascending model of pyelonephritis was established in Swiss Webster (LACA) female mice with these isolates. Quantitative bacterial count and histopathological evaluation of mouse renal tissue was done which were then assessed for a possible association with elaboration of virulence factors. RESULTS: All isolates of P. aeruginosa were able to colonize renal tissue of mice. However, renal counts varied amongst different isolates producing different virulence factors. Isolates producing high levels of haemolysin along with other virulence factors were able to colonize and multiply more in mouse renal tissue as compared to those producing low levels of haemolysin. INTERPRETATION & CONCLUSION: The findings of this study indicated an association between haemolysin production and renal colonization. High level of haemolysin production in vitro could be used as surrogate information for assessing pyelonephritic potential of P. aeruginosa.

Outbreak of cholera in & around Chandigarh during two successive years (2002, 2003).

BACKGROUND & OBJECTIVE: Outbreaks of cholera caused by Vibrio cholerae O1 Ogawa occurred in and around Chandigarh during two successive year 2002 and 2003. This study highlights the antibiotic sensitivity and phage typing pattern of V. cholerae isolates during 2002 and 2003. METHODS: Faecal specimens from acute gastroenteritis cases from July to September, 2002 and in the same month in 2003 were collected. Isolation and identification of pathogen was done according to standard methodology. Simultaneously water samples from the areas reporting the maximum number of cholera cases were also processed. Antibiotic susceptibility pattern of the isolates was studied and isolates were sent to National Institute of Cholera and Enteric Diseases (NICED), Kolkata for confirmation and phage typing. RESULTS: Of the 156 patients in 2002 and 125 in 2003, 59 and 40 isolates respectively were found to be positive for V. cholerae O1 serotype Ogawa biotype El tor. Of the 45 water samples tested in 2002, eight were found to be positive for V. cholerae O1 serotype Ogawa biotype El tor. None of the 52 water samples tested in 2003 was found to be positive for V. cholerae. Phage type 27 was found to be the predominant type for both the years. Majority of the clinical isolates were found to be resistant to more than two drugs. INTERPRETATION & CONCLUSION: The drug resistance in V. cholerae was on the rise during the subsequent outbreak. Phage 27 remained the predominant type in both the years. The major reason for the outbreak was traced to be contaminated water of the hand pumps in the affected area. Continuous surveillance of the outbreak is necessary to contain the spread of transmission.