Characterization of Potential Virulence Factors of Vibrio mimicus Isolated from Fishery Products and Water.

Vibrio mimicus is a Gram-negative bacterium that is closely related to V. cholerae and causes gastroenteritis in humans due to contaminated fish consumption and seafood. This bacterium was isolated and identified from 238 analyzed samples of sea water, oysters, and fish. Twenty strains were identified as V. mimicus according to amplification of the vmhA gene, which is useful as a marker of identification of the species. The production of lipases, proteases, and nucleases was detected; 45% of the strains were able to produce thermonucleases and 40% were capable of producing hydroxamate-type siderophores, and the fragment of the iuT gene was amplified in all of the V. mimicus strains. Seventy-five percent of V. mimicus strains showed cytopathic effect on Chinese hamster ovary (CHO) cells and destruction of the monolayer, and 100% of the strains were adherent on the HEp-2 cell line with an aggregative adherence pattern. The presence of virulence factors in V. mimicus strains obtained from fishery products suggests that another member of the Vibrio genus could represent a risk to the consumer due to production of different metabolites that allows it to subsist in the host.

Identification of capsule, biofilm, lateral flagellum, and type IV pili in Vibrio mimicus strains.

Vibrio mimicus is a bacterium that causes gastroenteritis; it is closely related to Vibrio cholerae, and can cause acute diarrhea like cholera- or dysentery-type diarrhea. It is distributed worldwide. Factors associated with virulence (such as hemolysins, enterotoxins, proteases, phospholipases, aerobactin, and hemagglutinin) have been identified; however, its pathogenicity mechanism is still unknown. In pathogenic Vibrio species such as V. cholerae, Vibrio. parahaemolyticus and Vibrio vulnificus, capsule, biofilms, lateral flagellum, and type IV pili are structures described as essential for pathogenicity. These structures had not been described in V. mimicus until this work. We used 20 V. mimicus strains isolated from water (6), oyster (9), and fish (5) samples and we were able to identify the capsule, biofilm, lateral flagellum, and type IV pili through phenotypic tests, electron microscopy, PCR, and sequencing. In all tested strains, we observed and identified the presence of capsular exopolysaccharide, biofilm formation in an in vitro model, as well as swarming, multiple flagellation, and pili. In addition, we identified homologous genes to those described in other bacteria of the genus in which these structures have been found. Identification of these structures in V. mimicus is a contribution to the biology of this organism and can help to reveal its pathogenic behavior.

Relationship of aquatic environmental factors with the abundance of Vibrio cholerae, Vibrio parahaemolyticus, Vibrio mimicus and Vibrio vulnificus in the coastal area of Guaymas, Sonora, Mexico.

Members of the genus Vibrio are common in aquatic environments. Among them are V. cholerae, V. vulnificus, V. parahaemolyticus and V. mimicus. Several studies have shown that environmental factors, such as temperature, salinity, and dissolved oxygen, are involved in their epidemiology. Therefore, the main objective of this study is to determine if there is a correlation between the presence/amount of V. cholerae, V, vulnificus, V. parahaemolyticus and V. mimicus and the environmental conditions of the seawater off the coast of Guaymas, México. Quantification of all four pathogenic bacteria was performed using the most probable number method, and suspected colonies were identified by polymerase chain reaction (PCR). Correlations were found using principal component analysis. V. parahaemolyticus was the most abundant and widely distributed bacteria, followed by V. vulnificus, V. mimicus and V. cholerae. Positive correlations between V. parahaemolyticus, V. vulnificus and V. mimicus with temperature, salinity, electric conductivity, and total dissolved solids were found. The abundance of V. cholerae was mainly affected by the sampling site and not by physicochemical parameters.

Listeria monocytogenes strains isolated from dry milk samples in Mexico: occurrence and antibiotic sensitivity.

Dry milk is a particular concern in Mexico, as approximately 150,000 metric tons of dry milk are imported every year at a cost of around $250 million. Dry milk is used to make many products, most of which are dairy products widely distributed among the population covered by welfare programs. The purpose of the study described in this article was to determine the presence of Listeria spp. in imported dry milk samples in Mexico, and to determine the sensitivity of the Listeria monocytogenes isolates to different antimicrobial agents. Listeria isolates (7.8% of 550 bacterial isolates) were identified as L. monocytogenes (53.49%), L. innocua (30.23%), L. seeligeri (13.95%), and L. ivanovii (2.33%). L. monocytogenes strains isolated showed multiresistance to ampicillin, erythromycin, tetracycline, dicloxacillin, and trimethoprim-sulfamethoxazole (9%-14%). The results provide additional evidence of the emergence of multiresistant Listeria strains both in nature and in widely consumed dairy products, representing a potential threat to human health.

Presence of virulence markers in environmental Vibrio vulnificus strains.

AIMS: This work aims to demonstrate the presence of several genes and factors associated with virulence in strains isolated from the environment at Pueblo Viejo Lagoon, State of Veracruz, Mexico. METHODS AND RESULTS: In this study, we investigated the production of V. vulnificus virulence factors, as cytolysin (haemolysin), RTX toxin, metalloprotease, siderophores, capsular polysaccharide, adhesion structures (like type IV pili), and polar and lateral flagella, involved in swimming and swarming (or, at least, the presence of genes encoding some of them) in 40 strains of V. vulnificus isolated from water and food. The results indicate that strains of environmental origin possess potential virulence characteristics. CONCLUSIONS: Caution should be exercised when consuming raw shellfish (especially by those more susceptible risk groups). SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first work focused on the evaluation of V. vulnificus virulence factors in Mexico.

Comparative studies on Enterococcus, Clostridium perfringens and Staphylococcus aureus as quality indicators in tropical seawater at a Pacific Mexican beach resort.

Three microorganisms were assayed to evaluate the microbiological quality in the seawater at a resort on the Mexican Pacific coast, and to test for possible associations among the titers of the various bacteria, their possible correlations with environmental conditions, and with the location of potential wastewater outflows. Significant microorganism levels were found (at Caletilla beach, Hornos beach, and Papagayo beach, respectively: for Enterococcus 157, 153, and 149, for C. perfringens 35, 89, and 56, for S. aureus 244,137, and 279CFU/100ml), often in excess of the presently set guideline values. In general, bacterial titers were higher during rainy season than in dry season. For S. aureus, in both seasons, highest concentrations were found at 3pm, the time of highest tourist presence at the beaches. Our results argue for the use of these three microorganisms as part of a set of indicators in the routine microbiological evaluation of Mexican beachwaters.

Occurrence and antibiotic sensitivity of Listeria monocytogenes strains isolated from oysters, fish, and estuarine water.

We analyzed the presence of Listeria spp. in oyster, fish, and seawater samples and tested isolates for antibiotic sensitivity. Listeria monocytogenes was found in 4.5% of fish samples and 8.3% of seawater samples and was not recovered from oysters. Multiresistant environmental strains were found, representing a potential threat to human health.

Isolation of Yersinia from raw meat (pork and chicken) and precooked meat (porcine tongues and sausages) collected from commercial establishments in Mexico City.

A total of 160 meat product samples were collected from commercial outlets in Mexico City to investigate the presence of different species of Yersinia by the 4 degrees C enrichment method after 1, 3, 5, and 7 days of incubation using alkaline treatment and isolating in cefsulodin-Irgasan-novobiocin and MacConkey agars with Tween 80. Overall, Yersinia spp. were isolated from 27% of the samples analyzed, whereas 40% of the raw and only 13% of the precooked samples were contaminated. Although 2,970 colonies showed Yersinia characteristics, only 706 (24%) actually corresponded to this genus: 49% were Yersinia enterocolitica, 25% Yersinia kristensenii, 15% Yersinia intermedia, 9% Yersinia frederiksenii, and 2% Yersinia aldovae; 10% corresponded to biotype 2, 2% to biotype 3, and 4% to biotype 4. The presence of Yersinia in raw and cooked meat products represents a health risk for consumers in Mexico, where further clinical studies are needed to assess the epidemiological importance of this pathogen.

Frequency of isolation of Campylobacter from roasted chicken samples from Mexico City.

The presence of Campylobacter spp. was investigated in 100 samples of roasted chicken tacos sold in well-established commercial outlets and semisettled street stands in Mexico City. From 600 colonies displaying Campylobacter morphology only 123 isolates were positive. From these isolates, 51 (41%) were identified as C. jejuni, 23 (19%) as C. coli, and 49 (40%) as other species of this genus. All of the 27 positive samples came from one location where handling practices allowed cross-contamination of the cooked product. The results indicate that these ready-to-consume products are contaminated with these bacteria, representing a potential risk for consumers, especially in establishments lacking adequate sanitary measures to prevent cross-contamination.

Prostate specific antigen and pathological features of prostate cancer in black and white patients: a comparative study based on radical prostatectomy specimens.

PURPOSE: We compared the relationship of pathological features and preoperative prostate specific antigen (PSA) levels of a consecutive series of black patients to a stage matched cohort of white patients treated during the same period. MATERIALS AND METHODS: The radical prostatectomy specimens of 40 black patients were reviewed and tumor volume was determined. Histopathological features (stage, grade, zonal distribution of cancer foci), tumor volume and prostate weight were correlated to pretreatment serum PSA levels. These parameters were compared with those of 148 white patients matched by pathological stage. RESULTS: Black patients exhibited a significantly higher incidence of seminal vesicle involvement (p=0.03) and cancers with a Gleason score of 8 or more (p=0.02), and a trend toward decreased pathologically organ confined, margin negative disease (40% black versus 53% white men, p=0.13). Although the incidences of multifocal cancer were virtually identical (90 and 82%) in the 2 groups, black patients exhibited a higher incidence of transition zone cancer foci (p <0.001). Mean prostate tumor volume, total gland weight and serum PSA level among black and white patients with pathological stage pT2-, pT2+ and pT3- cancer were not significantly different. However, with advancing pathological stage (pT3+ and pT3c) disease black patients had higher preoperative serum PSA levels on univariate and multivariate analyses despite similar total gland weight and tumor volume. CONCLUSIONS: Black patients who underwent radical prostatectomy often exhibited adverse pathological features. Two novel findings were that the distribution of cancer foci within the prostate was significantly different between black and white patients, and that serum PSA levels in patients with locally advanced prostate cancer were higher in black than in white men, despite adjustment for known variables affecting PSA. These observations suggest that differences in the biology of prostate cancer between these 2 races may exist.

Prostate cancer staging: correlation between ultrasound determined tumor contact length and pathologically confirmed extraprostatic extension.

PURPOSE: We determine whether a new parameter, the amount of tumor in contact with the fibromuscular rim (capsule) of the prostate, correlates with extraprostatic extension, and ascertain whether estimating the new parameter using transrectal ultrasonography can predict extraprostatic extension. MATERIALS AND METHODS: We analyzed step sectioned prostatectomy specimens from 189 patients who had had positive peripheral zone biopsies. We measured the contact length, maximum length (mm.) of the portion of the peripheral zone cancer that was in contact with the fibromuscular rim, and determined the contact ratio from the quotient (%) of the contact length divided by the tumor circumference. We evaluated the correlation between the pathological and ultrasound measurements of these parameters, as well as the accuracy of these criteria for predicting microscopic extraprostatic extension. RESULTS: Among the 189 cancers there was a significant difference (p <0.0001) between organ confined tumors and tumors with extraprostatic extension in contact length and contact ratio. There was a positive correlation (r = 0.691) between the contact lengths measured ultrasonically and histologically among 95 patients who had hypoechoic lesions associated with positive biopsies. A receiver operating characteristics curve of the ability of ultrasound estimated contact length to predict extraprostatic extension revealed the best cutoff value to be 23 mm. with 77% accuracy. Logistic regression analysis revealed that pathological contact length correlated better with extraprostatic extension than tumor volume, Gleason score, prostate specific antigen (PSA) level and pathological contact ratio. The best preoperative predictor of extraprostatic extension was the ultrasound contact length, followed by the contact ratio, PSA value, percentage of the biopsy specimen that was cancer and presence of perineural invasion in the biopsy specimen. Multiple logistic regression analysis revealed that the predictability of ultrasound contact length was improved by considering PSA value also. Probability plots for predicting extraprostatic extension were developed by combination of ultrasound contact length with PSA value. CONCLUSIONS: The length of tumor contact with the fibromuscular rim is more significantly related to extraprostatic extension than tumor volume, PSA level and tumor grade. For hypoechoic cancers a new ultrasound staging criterion, contact length, has been defined. For men who are clinically candidates for radical prostatectomy and have peripheral zone hypoechoic cancers the combination of ultrasound contact length and PSA value is the best predictor of extraprostatic extension.

Comparative analysis of prostate specific antigen and its indexes in the detection of prostate cancer.

PURPOSE: Prostate specific antigen (PSA) density and age referenced PSA have been proposed in an attempt to improve the power of PSA in the detection of early prostate cancer. Reported results have been controversial and disappointing. Because the association of gland volume with PSA is stronger than that of age we developed a new index, volume referenced PSA, and compared it to PSA density and other indexes. MATERIALS AND METHODS: Volume referenced PSA was developed from a control group of 408 men without clinical evidence of prostate cancer using a standardized Z score. A retrospective analysis was performed comparing PSA and all its indexes in 580 men who underwent prostate biopsy. In addition to overall analysis, PSA and its indexes were evaluated with receiver operating characteristic curves by age and volume subcategories. RESULTS: Cancer was identified in 35% of the 580 men. The number of missed cancers using established thresholds significantly favored volume referenced PSA clinically and statistically compared to all other indexes but it was equivalent to PSA alone clinically. Age referenced PSA prevented more biopsies from being done than did volume referenced PSA (39% versus 31%) but resulted in the diagnosis of 48% fewer cancers. Receiver operating characteristic curve analysis demonstrated a significantly better performance for volume referenced PSA and PSA density compared to PSA alone and age referenced PSA. CONCLUSIONS: Volume based PSA indexes are superior to PSA and age referenced PSA statistically. However, clinically volume referenced PSA is comparable to PSA, and both are superior to age referenced PSA and PSA density in the detection of prostate cancer.

Tumor volume and prostate specific antigen: implications for early detection and defining a window of curability.

PURPOSE: We attempted to determine the relationship between tumor volume and extent of localized prostate cancer, as well as the interrelationships of tumor volume with prostate specific antigen (PSA) level, grade and stage. MATERIALS AND METHODS: Serial whole mount sections from 128 patients who underwent radical prostatectomy were analyzed using a computer assisted volumetric program. Statistical evaluations were performed using logistic and simple regression analyses. RESULTS: The median tumor volume for patients with organ confined disease was significantly lower than for those with extraprostatic extension (1.25 versus 2.94 cc, p < 0.001). A significant incidence (32%) of small volume cancers (0.51 to 1.5 cc) exhibited extraprostatic extension while that of extraprostatic disease increased to 66% for patients with tumor volumes greater than 1.5 cc (p < 0.001). Of men with clinically significant (greater than 0.5 cc, or Gleason score 7 or more) pathological stage B disease 31% had a serum PSA value of 4 ng./ml. or less. Multivariate regression analysis of tumor volume as a function of PSA, grade and stage demonstrated that log PSA had the strongest association with tumor volume. Goodness-of-fit analysis (coefficient of determination) revealed that only 40 to 50% of the PSA levels are explained by tumor volume. CONCLUSIONS: These data suggest that the window of curability for prostate cancer decreases significantly once the tumor grows to a volume greater than 1.5 cc, and that grade and tumor volume are more significantly related to stage than PSA.

Management of transitional cell carcinoma involving von Brunn's nests.

We reviewed data collected from 371 patients with superficial transitional cell carcinoma of the bladder to determine whether carcinoma within von Brunn's nests is a risk factor for cancer progression and an indication for radical cystectomy. Cystectomy was done in 20 of 73 patients (27%) with transitional cell carcinoma in von Brunn's nests and in 42 of 298 patients (14%) without von Brunn's nest involvement. There was no significant difference in the Kaplan-Meier crude and disease-specific survival between patients with and without transitional cell carcinoma in von Brunn's nests, irrespective of whether radical cystectomy was performed initially. Of those patients with von Brunn's nest involvement none who underwent cystectomy died of bladder carcinoma, while 3 (6%) managed conservatively died of bladder cancer. Of those patients without von Brunn's nest involvement 1 (5%) managed with cystectomy and 9 (4%) managed conservatively died of bladder carcinoma. Furthermore, only 8 patients (15%) with and 29 (11%) without transitional cell carcinoma in von Brunn's nests showed disease progression after initial conservative management. Based on this analysis, our conclusion is that transitional cell carcinoma within von Brunn's nests is not a risk factor for disease progression or an absolute indication for radical cystectomy.

Diagnostic testing for prostate cancer detection: less is best.

The incidence of clinically organ-confined prostate cancer has markedly improved in programs designed to detect this disease by utilizing a combination of diagnostic modalities including digital rectal examination (DRE), transrectal ultrasonography (TRUS), and prostate-specific antigen (PSA). Biopsies were performed on 436 men who had abnormal findings on DRE, TRUS, or PSA. Overall, 39 percent of these men had histologic confirmation of prostate cancer. TRUS diagnosed more cancer (94%) than either DRE (80%) or PSA (89%), while the combination of TRUS and PSA diagnosed a similar number of cancers as the combination of DRE and PSA (100% vs 98%). The positive predictive value (PPV) of DRE and PSA combination was significantly better than that of TRUS and PSA (p = 0.01), but was not different from that of a combination of all three tests. DRE and PSA would have missed 4 cancers, but would have cost approximately 140 percent less than any program employing TRUS. Consequently, we recommend that the combination of DRE and PSA be used in primary early detection for prostate cancer and that TRUS be performed only when either or both DRE and PSA results are abnormal.

[Isolation of Campylobacter jejuni ATCC 29428 from inoculated fried pork meat and roasted chicken]. / Recuperación de Campylobacter jejuni ATCC 29428 a partir de carne de cerdo frita y de pollos rostizados inoculados.

The human gastroenteritis caused by Campylobacter jejuni in some industrialized countries is higher than gastroenteritis produced by Salmonella and Shigella. This has induced the development of techniques to demonstrate the presence of the microorganism in different foods using some culture media combinations. There is not a method to isolate C. jejuni from roasted chicken and fried pork meat, which are popular foods in México. The sensitivity of two culture media combinations was compared: Rama broth (RB)-Rama agar (RA) and Preston broth (PB)-Skirrow agar (SA) to isolate C. jejuni from these foods. The RB-RA combination demonstrated to be the best one to isolate C. jejuni.

The distribution of prostate specific antigen in men without clinical or pathological evidence of prostate cancer: relationship to gland volume and age.

We estimated the in vivo prostate gland volume in 408 men (320 without clinical evidence of prostate cancer, and 88 with an abnormal digital rectal examination and/or transrectal prostate ultrasound and negative biopsies) using sequential step-section ultrasound analysis and correlated it to the serum prostate specific antigen (PSA) value. Of the men 331 (81.1%) had a PSA value of 4 ng./ml. or less. The PSA value was greater than 4 but less than or equal to 10 in 64 men (15.7%) and greater than 10 in 13 (3.2%). The men were subclassified by prostate gland volume at arbitrary break points. A total of 139 men (34.1%) had a gland of 25 cm.3 or less, 2.2% of whom had a PSA value of greater than 4. Further analysis revealed that the incidence of a PSA value greater than 4 increased as the prostate volume increased (18.4% for greater than 25 but less than or equal to 50, and 65.4% for greater than 50) and as age increased. We found a statistically significant association between prostate gland volume and patient age (p less than 0.00005) to the serum PSA concentration. The finding of a PSA value of greater than 10 was uncommon regardless of the prostate gland volume. Clinical implications of these results are discussed, and a statistical model to estimate the serum PSA by gland volume and patient age was constructed.

Monoclonal prostate-specific antigen in untreated prostate cancer. Relationship to clinical stage and grade.

The authors evaluated 440 men with clinically staged and untreated prostate cancer with a monoclonal prostate-specific antigen (PSA) assay. The serum PSA value correlated significantly with both the stage and grade of disease (P less than 0.00005). The relationships between PSA and consecutive Stages A, B, C, and D2 (alpha = 0.15) and between progressive Gleason's scores 2 to 4, 5 to 7, and 8 to 10 (alpha = 0.15) were statistically significant. Also statistically significant was the correlation between serum PSA level and intracapsular versus extracapsular disease (P less than 0.00005), although no one value can be used to differentiate reliably between patients in these two categories. The probability of clinically detectable metastasis (Stage D2) is 85% if the serum PSA level is greater than 30; however, 12% of patients without clinical evidence of metastases (Stages A, B, and C) have such a serum PSA value. Despite the statistically significant association between PSA and tumor differentiation and volume as reflected by tumor grade and clinical stage, this marker cannot be used to determine either for an individual patient.

Early detection program for prostate cancer: results and identification of high-risk patient population.

Three hundred sixty-two men underwent transrectal ultrasound of the prostate (TRUS), digital rectal examination (DRE), and serum prostate-specific antigen (PSA) determination as part of an early detection program for prostate cancer. Thirty-seven (10%) cancers were detected. DRE had the highest sensitivity and specificity, 89 percent and 84 percent, respectively. TRUS and PSA had comparable sensitivities (84% and 81%) and specificities (82% and 82%). The positive predictive values of DRE, TRUS, and PSA determination were 39 percent, 35 percent, and 33 percent, respectively. We found a cancer detection rate of 16 percent among patients with symptoms of bladder outlet obstruction and 5 percent in patients without these symptoms. The detection rate was 36 percent for physician-referred patients and 3 percent for self-referred patients. This suggests to us that at the present time the best utilization of medical resources to increase prostate cancer detection is to educate men to have annual medical evaluations by primary-care physicians who are encouraged to incorporate risk assessment and screening DRE as part of their routine practice. Any man with either abnormal findings on examination or increased risk should be referred to a urologist for further evaluation.